Chapter

Biochemical Compounds
Tlis chapter is designed to haye the student thinh about the basic building blocks af biochewical
conrpounds, such as amino acids, which lead. to lsroteins; nitrogenous bases, which lead to nucleosides, mucleotides, and nucleic acids; and fatty acids, vlich lead to plmspholipids. The student will slso consider the biochemical function of intracellular c otltparttnentation in eukary otes, such as the nuclews, cndoplasmic reticulttnt, Golgi appdretus, ly sosome, ttritochondna, peroxisonte, dnd menrbranes. As fhis is a buildingbloch chapter, the references to dismse are sparse but will increase in later chapters of this
booh.

(A)
(B)

3 6

(c)

(D) 7 (E) 8
An Alrican native who rs going to college in the Untted
States experlences digestive problems (bloating, diarrhea. and flatulence) whenever she eats loods containing milk products. She is most likely deficlent in splitring whrch type of chemical bond?

(A) A sugar bond (B) An ester linkage

(C) A phosphodiester bond

QUESTIONS
Select the single best answer.

(D) An amide bond (E) A glycosidic bond

Consider the amino acld shown below. The configuration about r,vhich atom (labeled A through E) will derennine whether the amino acid rs in the D or L configurarion?

The procedure of Southern blottlng involr.es treatment of the solid support (nitrocellulose) containing the DNA u,ith NaOH to denature the double helix. Tieatment of a Northern blot r,vith NaOH, however, will lead to the hydrolysls of the nucleic acid on the fi1ter paper. This is due ro whi.h major chemical leature ol rhe nucleic et icls invoh,ed in a Northern blot? (A) The presence of thymine (B) The presence of uracil (C) The presence of a 2'-hydroxyl group (D) The presence ol a 3'-hydroxyl group (E) The presence of a 3'-5' phosphodiesrer hnkage -{ 6-month-o1d infant, with a histcry of chronic diarrhea and multlple pneumonras, is seen again by the pedra, trician for a possible episode of pneumonia. The chest \-ray shor,vs a pneumonia, but also reveals an abnorina11y sma11 thymus. Blood work shor,vs a distincr lack ..1 circulating iymphocytes. The most like1y inhented -:nzymatic defect in this child leads to an rnabrlity to -riter a purine nucleotide at which position of the ring
.ll-UClUre?

D/ -\

/rR-C-C-OA/
NHs*

r \r,i o/ ./c
/

Your patient has a mechanical heart valve and is chror-rica1ly anemic due to damage to red blood ce11s as rher pass through this r.alve. One of the signals that rarge: damaged red blood celis for removal from the circr-ri..,tion is the presence ol phosphatrdylserine in the outt: Ieaflet of the red ce11 membrane. Phosphaticirlsenn.-

is an integral part of cell membranes and is nonlr:l '. lound in the inner leaflet of the red ceLl membran: fhis flip-flop of phosphatidvlserine berr,veen me n-Lhr.,.::-:

phosphatidylserine to ieaflets exposes which part of the 'the enl'ironment?

(A) The head grouP (B) FattY acids (C) SPhlngoslne (D) Glycerol

A rype I diabetic is brought to the . emergency ;;p;;;.;, due to lethargv utJ rapidbreathing Blood *.urur.-..ts indicated eit'ur"d levels of glucose and patient was ketone bodies. Blood pH was 7 1 The
the folloirlng
blood pH? (A) oxygen
gases

which one of ."frrUur"g enhanced breathing to exhale

in

order

1o correct the abnormai

(E) Ceramide

6:', Whrch of the followi'ng is the type^ of bond '" "t' ,-r.,.1.otid es [o form long pollT rrers?

that

a11ows

(B) Nitrogen (C) Nitrous oxide , (D) Carbon dioxide

OH lll R_C-N-R'
o

(E) Superoxide
of the^pH in the The proteln albumin is a major bufler and 7 4' blooi, which is normally kept between 7'2 slde chai'n of Which of the followtttg i' u'-' n*ino acid range? albumin that participates in this buffering

R-c-o-R'
o

il

R-o-P-o-R',
oI

II

(A) Histldine (B) Aspartate (C) Glutamate (D) Lysine (E) Arglnine

R-c-o-P-o-R'
oI

illl

.,{ii

Consider the following s[ructure:

R-o-R'
A couple has had five children'
of who exhlbit short muscle weakeyelid droop, and some degree of (some severe' some mild) The
a-li

HoHHoHHgHHoI l.! I I ll 'l'ii I I ii t HoN* -C -c - N' -c -c -* -q -c -ttt -c -c -o,,3'r " -

Y H

,ru,rr.,

i I CHzOH CHs ?" cooI

,-r.s. urla'hearing ltss at a mild 1eve1' ,rrott-t., also has Jrch problems, although mutation that afflicts The father has no qrmptoms' The *ort liktly resides in DNA found in which
ii-r"

This structure is
following? (A) An amino acid (B) A triPePtlde (C) A tetraPePtide (D) A lipid (E) A carbohydrate

best descrlbed

as which of the

Jrfa*"

i.ntracellular organelle?

'

14) Mitochondria

(B) Peroxisome (C) Lysosome (D) EndoPlasmic reti'culum (E) Nucleus

between 4 Lysosomal enzFnes have a pH optimum are kept at this pH ur-ra O. rn. intiaiysosomal contents b-v *-hich of the following mechamsms? (A) The actlve pumping of protons out of the organelle (B) The free d'iffusion of protons out of the organelle (C) The actiYe pumping of protons into the organelle organelle The lree diffusion of protons into the

weak base wrth A drug contalns one ionizable group' a free , ,i'"f 9.0. The d'rug enters cells viaform diffusion This will its uncharged ili".i*f-t-,fr. *embrane"ln ;:;;?"" readiiy at which of the foliowing pH vaiues?

(A) 3.5 (B) 5.5

(c)

7.0

lD)

it, ,fr.

lysosome srrlthesis of carboxylic acids within the

(D) 7.6 (E) e.)

I
I

BiochemicalCompounds 3

l3

Consider the five functional groups shor,vn below

this patient are most likely derived from whrch type ol

molecule?

(i)

NH2

(A) Purines (B) P1-rimidines (C) \rcotrnamides

(D) -\mino acids

(ii)
(iii) (iv)

(E) Fattl'

acids

"'(o OH
oH

t7

A single-stranded D\-\ nrolecuIe contarns 20o/aA,25okT, 30%G, and 259oC \\-hen the con.rplement ol this strand rs sl,nthesized, the T content of the resuitrng duplex r'vill be lvhrch one of the fo11ou'ing?

cH3

(A) 20% (B) 22.5"/"

(c)

25%
5a/o

(D) 27

(v) R-c=cHz
t8
A hydrogen bond would form between lvhich parr of
groups?

(E) 30%
The activated form of the drug omeprazole (used [o treal peptic uicer disease) prevents acid secretion by formlng a covalent bond with the H*, K*-AfPase, thereby inhiblting the enzyme's transport capabilities. Analysis ol the

(A) ilr and iv

(B) rC) (D) (E)

iii and v
ri and iv

drug-treated protein demonstrated that an internal

cysteine residue was involved in lhe covalent interaclion r,vith the drug. Further analysls indicated that the bond was not susceptible to acid or base catalyzed hydrolysrs. Based on this lnformatron, one u.ou1d expect the drug to contarn which of the following functional groups that r,vould be critical for its inhibrtor\. aclion?

ii and iu i and v

l+ \Vater is the universal solvent lor biologtcai systems.

Compared to ethanol, for example, water has a reiatively hrgh borling point and high freezing point. This is due primarily to which one of the following properties of
$'ater?

(A) A carboxllic acid (B) A lree primary amrno group

(C) An imidazole group (D) A reactive sulfh1-dry'l group

,,{) 1ts hydrophobic effect ,B) lonic interactions between

,C) The pH

(E) A phosphate group

water moiecules

,.D) Hydrogen bonds between water molecules L Van der Waals rnteractions

r9 Your drabeiic patient has a hemoglobin Alc (HbA1c) ol 8 8. HbAIc dilfers lrom unmodified hemoglobin by
u-hrch one ol the lol1or'ving?

(A) Amrno acid

sequence

t5 \lembrane formation occurs, in part,

1

due to lor'v irpid sol-

(B)
(D)

Senne acylation
Intrace11u1ar

:brlity rn water due to pnmarily w'hich of the follor,irng? \) Hydrogen bond fomation betr,r,-een lipids and water
1
1

(C) Vaiine glycosylation

location

Bl

Covalent bond formation belr,veen hprds and rvater

(E)

Rate of degradatron

C) A decrease in water entropy Dl An increase in water entropy Er lonic bond formation betr'veen lipids and

r,valer

l5

-7-r-ear-old woman visits the emergency department to ser.ere pain rn the metatarsophalangeal (MTP) ----rt ol her rlght great toe. Upon examination, the toe . bright red, sr,vollen, warm, and very sensrlive to :: rouch. Analysis of ;oint fluid shou's crystals. The :,..---.nt is given indomethacin to reduce the severity of ..-. s\nptoms. The crystals that are accumulating in
---

ile

Lir.er catabolism of xenobiotic compounds, such as acetaminophen (Tyleno1), is geared toward increasing the solubility of such compounds for safe excretion from the body This can occur via the addition of which compound beiow'in a cor.,alent linkage with the xenobiotic?

(A) Phenyialanine (B) Palmitate (C) Linoleate (D) Glucuronate (E) Cholesterol

ANSWERS
1
The answer is (: The presence of a 2'-hydroxyl group. RNA is susceptible to alkaline hr-droh-sis. uhereas D\-1 is not. The malor dilference betu-een the nro polrrLucleotides is the presence ol a 2'-hr drorr I lrtrup rrll r-1e sugar ribose rn R\-{. \ arsus 1tj aL,.enit tl ,fe ..:'rlilasa. a component oi D\\ Ur:i:: ilk:.r-ne l-.r:ir:i:n.. ::.c h; drorr i qr.lup ,:: :. . -,: ; :.^- l. : -' r :- j -, .,-^ the phosphodiester iin^..ge t.:'.'...:r --.*--1..:-: :-.,.- iotides, breakrng the tn^:ge ;:; -e,'.:::: --- - . ::--.:-'-::--

the nucleoside inosine. The same type of reactlon occurs

in IRNA anticodons, in which a 5' position adenine is

converted to hlpoxanthine, to produce the nucleoslde inosrne. inosine is a wobble base palr former, having the ability to base pair with adenine, uracil, or cytosine.

is E: A glycosidic bond. The patienr is exhibiting the classic signs of lactose intolerance, in
The answer

formatron of a cychc nrcieoiide. ns ints !i:t aaa;: ;i every phosphodiester linkage in R\-{. hr drol,vsrs of the RNA will occur due to ihese reactions. As DNA lacks the 2'-hydroxyl group. this reaction cannot occur, and DNA is very stable under alkaline conditions. The fact that DNA contains thyrnrne, and RNA uracil (both true statements) does not address the base stability of DNA as compared to R\A. Both DNA and RNA contain 3'-hydroxyl groups, rvhich are usually in 3'-5' phosphodiester bonds ln the DNA backbone. The procedure of Southern blotting is used in the diagnosis of various dlsorders, including some instances of hemoglobinopathies and drseases induced by tdplet-repeat expansions of DNA (such as m,votonic dystrophy).

u'hich intestlnal lactase levels are 1ow, and the major dietary component of miik products (lactose) cannot be drgested. Lactase wili split the B-1,4 llnkage between galactose and glucose in lactose. The lactose thus
passes

unmetabolized to the bacteria inhabiting the gut,

and their metabolism of the disaccharide ieads to the observed symptoms. Combining two sugars in a dehydration reaction creates a glycosidic bond Adding a
sugar to the nitrogen of a nitrogenous base also creates an N-glycosidic bond. A sugar bond is not an applicable term in biochemistry Ester linkages contain an oxygen

Iinked to a carbonyl group. A phosphodiester bond is

a phosphate in two ester linkages with two different compounds (such as the 3'-5' Iink in the sugar phosphate backbone of DNA and RNA). An amide bond is the joining of an amino group with a carboxylic acid with the loss of water. These types of bonds are shown
below.

le

Th" answer is C: 6. The child is exhibiting the syrnptoms of adenosine deaminase deficiency, an inherited immunodeflciency slndrome that ls a cause of severe combined rmmunodeficrenclt The disease ts caused by the lack of adenosine deaminase (a gene found on chromosome 20), which converts adenosine to lnosine (part of the salvage and degradatrve pathway of adenosine, see ihe figurebelow). This dtsorder leads to an accumulation o[ deoxyadenosine and S-adenosylhomocysteine, which are toxic to immature l1-rnphocytes in the thymus. As indicated in the figure below, the amino group at position 6 ls deaminated and is replaced by a doublebond oxygen, to produce the base hJpoxanthine, and

B-C-O-R'
Ester linkage

il

o illl R-O-P-O-R', I oPhosphodiester bond

oH
R-C-N-R',
Amide bond
l

t
A B-glycosidic bond, which is cleaved by lactase

:q[
NHr

Numbering of the purine ring

(x)
R

t-

o
NHg

--^,J(-*.
R

R = Ribose

(-At'

Adenosine

D. The central (or tx) carbon of amino acids has four different substltuents (as long as R ls not H, in which case the amino acid is glycine). Due to havlng four dlfferent substituents, this is considered an as)..rnmetric carbon, and the orientation of the substituents around this carbon can be in either the D or L configuratlon. None of the other choices reler to an asymmetric carbon atom. Many biochemlcal compounds (including drugs) are only active as either the D or L isomer. Fenfluramine, an appetite suppressant, in only active in its D form; in its L form it induces
The answer is drowsiness.

Adenosine deaminase reaction

Biochemrcal Cr-.:-:_

-.',: ;

The answer is A: The head group. phospholipids :,-ntain a very hydrophobic backbone and a ,,head ::.rup" that is primarily hydrophilic. The hydrophobic :.rrtion of the phospholtpid remarns embedded in the
::embrane whl1e the hydrophilic head group faces the ol the cell As seen in the figure :elou', the glycerol portlon (or ceramide portion, which .Lrnrains sphrngosine) of the phospholipid, as well as the attr- acrds, remains embedded in the membrane while ..nli-the head group (R) laces the aqueous environment. Thus, rvhen a phosphohpid flip-flops across rhe mem_ lrane, the head group will always end up facing the aqueous environment.
j-.queous environment
Aqueous phase
H R

muscular dystrophy, includrng Kearns-Sar.e : ...:_. (this case), Lelgh sl,ndrome (non-X-hnkeo i.r-,:.

sprdrome, mitochondrial DNA depletion and mitochondrial encephalomyopath;r

s\:.:- -

-_

hydrolysis provides the energy to pump prorons agains: their concentration gradient. The removal of protons from the lysosome l,vould raise pH, not lower it (therebr

The ansurer is C: The active pumping of protons into the organelle. Lysosomal membranes contain an en_\:t:c which actively pumps protons into the organal-:. thereby maintaining a low intraorganelle pH t,. : enzlrne is the proton-translocating Afpase, as -{TF

HO

a\ /--\ /- \

) )\_) iiili: itil;: :iiii: litiil

II

rendenng answers A and B incorrect). Free ditfusron of protons would not a1low uptake of protons againsr a concentration gradient, as diffusion is the flow from a hlgher concentration to a lorver concentration. Since the cyroplasmic pH is in the range ol 7 .2, if protons were freely dillusible across the lysosomal membrane, the protons rvould leave the lysosomes and enter the
c) top1asm. The lysosomes do not sy,r,rthesize large amounrs ol carboxylic acids (a neak acid) in order io lorver the pH instde the organelle.

lnterior of membrane bilayer

The answer is C. A phosphodiesrer bond links nucleotides in nucleic acids. Answer A rs an amlde bond (the

type found llnking amino acids together in protelns). Answer B is an ester linkage (the type found in tria_
cy1glycero1, in w.hich fatty acids are attached to a glyc_ erol backbone). Answer D is a phosphoanhydrlde bond (similar ro rhar found at the l position of 1,3 bisphos_ phoglycerate), and answer E is an ether linkage (found ln ether lipids, for example).

The answer is A: Mitochondria. The mother and chil_

dren are experiencing the effects of a mitochondrial disorder. Eukaryotic cells actually have two genomes; one in the nucleus, and another in the mrtochondria. The mitochondrial genome codes for a small number of proteins which are found in the mitochondrla. In order lo make these proteins the mitochondria also sl,nthesize their own IRNA molecules. As only the mother transmits mitochondria to her children, mitochondrial
eases dis_

The answer is D: Carbon dioxide. The patient is in the midst of diabetic ketoacldosls, tn whichihe procluction, but nonuse, of ketone bodies (which are acids) results in a srgnificant lowering of blood pH This patienr will be creating a respiratory aikalosis to attempr to com_ pensate for a metabolic acidosis. Under conditions of an acidosis, the proton concentration of the blood needs to be reduced. Due to the presence of carbonrc anhy_ drase in the red blood ce1l, as carbon dioxide is exhaled, protons are removed from solution. As the concentra_ tion of carbon dioxide is reduced, bicarbonate (HCO- ) reacrs wirh a proron (Ht) to form carbonic acid, whiih then dissociates to form water (H,O) and carbon cliox_ ide (COr). These reactiorr, ,.. ,r_,rr*arized in the figure below. Thus, as carbon dioxide is exhaled, the proton concentration decreases, and the acidosis is reduced. The exhalation of oxygen or nitrogen will not affect the proton 1evels in the blood, nor will the loss of nitrous oxide or superoxide.

display a unique inheritance pattern. None of the other organelles listed, other than the nucleus, conrain DNA, and these sy.rnptoms and inheritance pattern are not consistent with a mutation in nuclear DNA. The mitochondrial genome is 15,569 base pairs in size, encoding 37 genes. These genes include two differ_ ent molecules of rRNA, 22 different IRNA molecules. and 13 pollpeptides (seven subunits ol NADH dehydrogenase, or complex I, three subunits of cytochrome c oxidase, or complex I! two subunits of the proton translocating AfP sprthase, and cytochrome b). There are multiple mitochondrial disorders associared with

CO2

11rg

-+

H2CO3

=::-

H+

HCOS-

As the concentration of carbon dioxide decreases, the equilibrium is shifted to the left, thereby also decreasing the proton concentration, resulting in a rise in pH.

t0

The answer is A: Histidine. Of the amino acid choice s listed only histidine has a side chain r,vhich could ccr_ ceivably buffer in rhe range of 7 .Z to 7..1. The in.r1;-.._ zole group of histidine has a pK. of 6.0, but rhis c::: :: altered by the local environmenr of the prtrLein _\s::.:.- acid and gluramic acid have side chain carbo\-, 1,. . - - .

Chapter

each ol u'hrch has a pK, about '1.0 and would not be able to contnbute to duffering at neutral pH Both

14 "

The ansurer

lysrne and. arginine have basic side chains, wrth pK, .lalues about 9.5, and those too will not be able to buffer
near neutral PH.

bonds between water molecules. Water exhibits its unique properties d-i-: to the extensive hydrogen bonding that can occ-: between water molecules, and due to the extreme'-' high concentration of water (at lSg/mo1, I I- of r'r-a:''
contains 55 moles of water, for a concentration apprt')-imating 55M). Water forms hydrogen bonds in a i::"tlcelike structure (see the figure below), which make= "

is D: Hydrogen

ll

The answer is C: A tetrapeptide. The s[ructure consists of four amino acids linked by three peptide bonds, gen-

erating a tetrapeptlde (the amino acids are glycine, serine, alanine, and aspartic acid). The structure con[ains no lipid or carbohYdrate.

t?

The answer is E: 9.2. With a pK of 9 0, the weak base needs to lose a proton to enter cel1s in its uncharged form (this base is mos[ likely -NHr* below pH 9 0, and -NH,

difficult for water to leave and become gaseous (thus il' high boiling point). As waler moYement rs reduced ci": to 1ow temperature, the lattice becomes a solid (ice explaining the relatively high freezing point o[ wat;: fhe hydrophoblc effect of water comes into play ll'hen : hydrophobic substance enters water; it does not applr --water itself. The concentration of water molecules, s-t--7\1 i-' a charge, is very small (at pH 7.0, there is I x I0

above pH 9.0). Thus, the higher the pH, the greater the proportion of drug which rs in its unionized form At pH uulr., 1.r, than 9.0, greater than 50o/o of the drug will be ionized, which will slow its entry into cells At pH 9 '2, less than 50% of the drug is ronized, and as the umonized form enters the ce1l, lt will reduce the concentralion

of unionized drug rn the circulation, thereby forcing a re-equilibration and generatlng more unionized drug At
the next highest pH value listed, 7 6, less than
8o/o

and OH- ions, out of a 55 M solutlon), so lonic inter;':tions between water molecules are minimal and do n-contribute to rts high boiling and freezing points T:' pH of the water refers to the concentration of proto::' which wr11 not affect the hydrogen bonding capacitr ' the water molecules. Van der Waals interactions do r-: play a role in the physical properties of water'

of the

drug is unionized, and the rate ol transport would be much less than at PH 9.2.
Hydrogen bonds are formed atoms share a hydrogen' The when two electronega[ive atom which has a greater affinity for the hydrogen is known as the hydrogen bond donor, and the atom with
The answer is D: ii and

13

iii.

the lesser aff,nity the hydrogen bond acceptor' Hydrogens linked to carbons never partrcipate in hydrogen tonding, as the electrons in the bond are evenly shared by the hydrogen and the carbon. In the case of hydrogens bound to nitrogen, or oxygen, the electronegative Ito* hus a higher afflnity for the electrons, thereby

allowing hydrogen to "bond" to another electronega-

t5-

tive atom. Of the structures shown, structure i could be a hydrogen bond acceptor or donor, and the carbonyl group of structure ir could be a hydrogen bond acceptr. The hyciroxyl group in structure iii can either be a clonor or the oxygen can be an acceptor' Compounds iv and v will not partlcipate in hydrogen bonding due to containi.ng exclusively C-H bonds. Thus, of the choices Iisted, onLy compounds ri and iii would form a hydrogen bond, as indicated below.

The answer is D: An increase in water entropy' Lipids are hydrophoblc molecules which do not form hydrogen bonds with r,vater. Due to this, water molecules wrl1 io.* u "cage" around the lipid molecules, surroundlng

R-o
*

/H'---_..-_-

d* -"'-- HO ,C-^',

them. Cage forming decreases water entropy, which is unfavorable, and this leads to the hydrophobrc effect, in which the lipld molecules all come together such that only one large cage needs to be formed about the lipid molecules, rather than many sma11 cages about each inclividual lipid mo1ecu1e. The lipids do not form covalent or ionic bonds with water, and, as mentioned above, lipids in water leads to a decrease in water entropy (which is unfavorable), rather than an increase

in the entropy of water (which would be a favorable

event). The figure below shows a "cage" of water surrounding ten llpld molecules.

= Hydrogen bond acceptor

Biochemical

Com:,-,^::-.

/H -H-----o\ -----H

-H.

duplex. The [A] in the duplex rl,r11 also be 22 i:: .,=since [A] = [Tl), and the concentrations ol lG :,:-: will each be ).7 .5ok for the duplex.

Hu

(cH3(cH2)ncH3ho

Hs
t6

)o---- n-o'-r------.b/y

,H

"o-H I

18

The answer is D: A reactive sulfhydryl

group. A i:.:

The answer is A:
a

Purines. The patient is sulfering from

gout attack due to the buildup of urlc acld in the b1ood, and precipitation of uric acid in "cold" areas of the bod1,, such as the great toe. Uric acid has the basic rlng structure of the purines and is the degradative product of adenine and guanlne. As shown in the figure belou', the ring structure of uric acid is not at all srmilar to p1'rin.ridines, nicotinamides (derlved from the r.itamin niacin), amino acids, or fatty aclds.

hydrolysis. Forming a bond with the other groups hsted would lead to relatively easy hydrolysls reactions, rendering the rnhibitory bond unstable Since the inhibrtion is stable, the best choice rs a sulfhydryl group. The drug is a proton pump inhibitor and reduces acid secretion by the chief ce1ls in the stomach, thereby alleviatrng symptoms of acid reflux in the patient.
19

sulfhydryl group ln the drug would be able tc, lonr a disulfide bond with the protein (-CHr-S-S-CH' which is an oxidation reduction reaciion. This ri ouil render the drsulfide resistant to acid or base-catalr -ei

t,H.{ t,Ho }^t, -

^,, .^,, , ^z.O :6-

General structure of a fatty acid

T-Y\ \*A*' n \r/

o
il

Generatstructure
ot a Purrne

Generatstructure
of a Pvrimidine

glycosylation. HbAlc ls glycosylated hemoglobin, reflectlng the 1eve1 of blood g1ucose over the lifetime of the erythrocyte (120 days). The higher the concentration o[ HbAlc, the more poorly controlled blood glucose levels are (normal is about 5.5olo HbAlc). The glycosylation prlmarily occurs on the N-terminal valine residues of the B chains (which contain a free amino group). The amino acid sequences of hemoglobin and HbAlc are the same, there is no fatty acid addition (acylation) to the hemoglobin, the red cell contains no intracellular organelles for compartmentation to be an issue, and the rate of degradation of nonmodified hemoglobin and HbAlc are the same.
The answer is C: Valine
The answer is D:

r'>(" \*/
U

^_Ntu '""2 General

structure

ot nrcotrnamrde

Uric acid

20

Glucuronate. In order to make

xenobi-

^ .1. ^/: h-u-u--\o_ i

NHs+

Generat structure of an amino acid

t7

The answer is B: 22.51o. The given strand of DNA contains 25o/oT , the complementary strand will contain 20oloT (this must be equivalent to the content of A in the given strand, since A and T base pair, and tAl = tTl in duplex DNA). For the entire duplex then, the T content ls the average of 25o/o and 20ak, or 22.5ok lor the

otic more soluble, a hydrophilic group needs to be added to the xenobiotlc. Of the possible answer choices, only glucuronic acid (glucose with a carboxylic acid at posltlon 6 instead of an alcohol group) is a hydrophilic molecule. Glucuronic acid is added to the xenobiotic at position 1, using the activated intermediate UDPglucuronate. Once added to the xenobiotic, the highiy soluble glucuronate confers enhanced solubllity to the adduct. Phenylalanine contains a hydrophobic srde chain, and palmitaie, linoleate, and cholesterol are all very hydrophobic mo1ecules. Their addition to a xenobiotic would decrease. rather than increase, its solubility

Chapter 2

Protein Structure and Function

ln this chapter, questions will cover various
aspects of protein structure andfunction, including enzyme ltinetics, the transport of molecules across (A) A surface-associated domain
18) A very hydrophobic environment (C) A very polar environment (D) Buried deep within the core of this giobular protein

cell membrdnes, vnrious mechanisms of catalysis, the binding of oxygen to hemoglobin, and various diseases that result from altered protein folding. There will be a mixture oJ clinical and nonclinical questions seen in this chaPter.

(E) Surrounded by phenyialanine, valine, and leuclne

residues

QUESTIONS
Select the single best answer.

?j. A major driing force {or protein tolding is the hydro' phobic effect, ln whlch hydrophobic amino acid side chains tend to cluster together, usually in the core of
globular proteins. This occurs primarily due to which of the following? (A) Increasing hydrogen bond formatlon (B) Increasing the entropy of water (C) Increasing disulfrde bond formation

,'* A kinetic analysis of the effect of a drug on an enzymeb " '" activity was performed, and the results shown below
were obtained. The drug would be best classified which one of the following?
as

(D) Minimizing van der Waals i.nteractions (E) Reducing steric hindrance between amino acid side
chains

A 7-year-old Afrlcan American male is admitted to the hospital with severe abdominal pain. A blood workup
indicated anemia, and an abnormal blood smear (see below). The molecular event triggering this disease is whlch of the following?

)
14Sl

q-\) A competitive inhibitor (B) ,\ noncompetitive inhibltor tC) -{n uncompetitive inhibitor .rm.e 1D) -{ competitive activator of the er:z enzyme (E) -\ noncompetitive activator of the

A cniical htstidine slde chain in an en4'me's active site displal s a pK value of 8.2. Which of the following best descrites the-1oca1 enrironment in which this histidine
resldue resides?

Protein Structure and

F-::,-

--

(A) A ioss of quaternary structure of the hemoglobin

moiecule

(B) An

rncrease in oxygen binding to hemoglobln (C) A gain of ionic interactions, stabilizing the "T" form

Whiie working an overnight shifi in the e:::.::.--.'ir , '.'. department you are called to see an B-year-old appears to have a fracture in his arm. Upon ta.--:.. -,
history, you learn that this child has been to the ER::-^--

of hemoglobin

(D) An increase in hydrophobic interactions between

deoxyhemoglobin molecules

(E) An alteration in hemoglobin secondary structure

leading to loss of the "cr" helix A 56-year-o1d pathologist was taken to his family doctor by hls son for he was showing mood changes, minor loss of memory, and decreased motor skills During the patient history, it became clear that over the course of his career he had, on occasion, cut himself using the instruments he had been using on the cadavers he had been working on. A potential explanatlon for his symptoms is abnormal aggregation of which of the foliowing
protelns?

tiple times lor fractures, and the incldents that leac :-the fracture would be described as mlld trauma ar 'b.s: X-rays indicate a number of healed fractures that ia. boy and his parents were unaware of (see example ..: arm X-ray below). Physical exam shows sky blue scler: The parents then inform you that the child is takinE bisphosphonates for his condition. The mechanisn whereby the frequency of fractures is being reduced in this patient is which of the following?

(A) Hemoglobln ln the red blood cells (B) Fibrillin ln the extracellular compartments of the
brain

(C) A truncated neuronal protein (D) A misfolded form of a normal protein

(E) A truncated extracellular protein

A teenager, new to your practice, comes in for a routine
physical exam. His family had just moved to the city, and the boy had rarely seen a doctor before. Upon examination, you notice a high, arched palate, disproportionately long arrns and fingers, a sunken chest, and mild scollosis. The patient has been complaining of lack of breath while doing routine chores, and upon listening to his heart, you detect an aofiic regurgltatlon murmur. Careful examination of the eyes is indicated by the figure below. Based on your physical exam and history you are suspicious of an inbom error of metabolism ln which of the following proteins?

(A) Collagen rB) Fibrillin (C) Elastin (D) Dystrophin iE) B-catenin

(A) Increased slnthesis of collagen (B) Increased resorption of collagen (C) Decreased sgrthesis of collagen (D) Decreased resorption of collagen (E) lncreased rynthesis of fibri11in
You are visited by a 4}-year-oid female patient c.'::plaining of weight 1oss, numbness in the hand. ..::

t-

l0

Chapter 2
feet, lartgue , and difficulty swallowing. physical exam notes an enlarged tongue, enlarged liver, a rubbery feeling around the loints, and brulsing around the

e;-es.

bone marrow blopsy shows an abnormal

staining

of

denarured prorein (see below). These

denatured proteins are most Iikeiy to be which of the following?

(A) Creating lysine cross-links in collagen (B) Mobllization of calcium into bone (C) Hydroxylation of proline resiciues in collagen (D) Glycosylation o[ fibri11ln (E) Con,,,ersion of glycine to proline in coliagen

1!

A patient, who was recenrly diagnosed with cystic fibro, sis, displays an increased blood clotting rime. This is mosr likely due to which of the following? (A) Lack of proline hydroxylation (B) Inability to catalyze rransaminarions

(C) Lack of dolichol and an inability to glycosylate

serum protetns

(D) lnability to carboxylate glutamlc acld side chains (E) Reducrion in the synthesls of blood clotring facrors
due to lack of liprds for energy production

12

You order a hemoglobin electrophoresis on a parlcnl suspected of having sickle cell disease. A blood sample
was obtained and the red cells were isolated. Disruption of the red ce11s released the hemoglobin, which *ui ,.,r,

(A) Antibody iight chains

(B) Collagen

(c)

Flbrl11in

(D) Albumin

on a polyacrylamide ge1. Following the electrophoresis. a Western blot was performed to locate the hemoglo_ bln. The results of the Western blot are shown belou-. Which one of the following statements best represents the inrerpretation of the results?

(E) Tiansaminases

A family of four from New Jersey has embarked on a vacation in the Rocky Mountains. A11 four required a24 to 48h acclimation to the high altirude, as all were breathing at a rapid pace unril the acclimation rook
cells
effect. In addition to increasing the number of red blooci in circulation, what other compensatory mecha-

o
x

nlsm occurred within the red blood cell during this

acclimation period?

(A) Increased slmrhesis of lacric acid (B) Decreased s1,r-rthesis of lactrc acid (C) Increased s1,r-rthesis of 2,3-bisphosphoglycerate (D) Decreased slnthesis of 2,3-bisphosphoglycerare (E) Decreased degradation of bilirubin, producing
carbon monoxide

o
(A)
less

The band marked as X refers ro rhe wild-r)?e hemo-

globin protein

(B) The band in lane 2 represenrs an individual witl_

sickle
ce11 disease.

t0 ln the

1800s, British sailors on long seajourneys developed sore and bleeding gums, sometimes to the point

(C) A carrier of sickle

ce1l disease is represented

by thr
ce

that their teerh would loosen and fall out. The inrroduction of limes to their diets helped to prevent these
occurrences. The biochemtcal step that was lacking in
these sailors was which of the following?

band in lane 3 (D) Lane 1 represents an individual

disease

with sickle

(E) Lane 3 represenrs an individual with sickie

disease

ce

protein Stmcture and F,t:_--..

ll
_

t3

Shown be1ou, is a secrion of a protein r,vhich forms a tlpical s-heli-x In the form of an o,,heli_x, a hydrogen bond would be formed berr.veen which two of the iabel-ed aroms?

-!iiii!?iinir??i?f
@(o\2'
(A) A and C (B) B and D (C) B and E (D) B and F (E) D and F

ttll

@o@@
sH

trr
-.1.

t 2 unirs/mrn/mg prurein lB) -i unirs/min/mg prorein

(A

min/mg protein, wrrh a K, of I .25 prNl in rhe -,:.:: inhibiror, but in the p..r.,-,.. of 5pN,{ inhibii..: -:-.. is 6 units/min/mg protein, with the same K . ri i.r-, .. reloeitl olrh( reecl ion in rhe prcsence ol ;pt\l . : at a substrate concentration ol 2.50pM?

(C) o untrs/min/mg prorein

_ -c -N-c -c N_c _c _N_c _c _rrr_i_c _ru_c_c_ru_ ",r, cH: !r, ?n, ir.

./,/o
SH

(E) I0 unirs/rnin/mg
17

(D r 8 unirs/min/mg prorein

prorctn

child has trouble nsrng from a sitting iosition. Examina tron reveals calf l-rypertrophy and hmb,girdle r,veakness. The inborn error in this patient is due to which of the lollou,ing?

A 3-;,s2r-.16 boy is evaluatecl by the pecliatrician as rhe

(A) Defective muscle mitochondria (B) A mutation in rhe ol hemoglobin

(C) A delect in rhe srructure of the hepatoqte membrane (D) A defect in the structure ol the sarcolemma (E) A defect in the transcnptron of muscle_specifre gencs
B-chain

l4 A 37-year-o1d female has rrouble keeping her eyes open and swallowing and is beginning ro ,t.r. her speech
The patient has also noticed u -.rk."r. in her arms and iegs. Tieatment mth edrophonium chlorjde resujts rn a temporary relief of symproms. The untlerlying etiology

t8

oI this disorder involves auto-antibodtes that do r,vhich ol the iollowrng?

(A

plays a microcytic anemia rvith Jnormal erythrocytes (see picture belolrl under al1 conclitions. this detect is most likel),due ro a hereditary murarion in which of the
lollorvrng?

An S-monrh-old infanr exhibirs.laundlce ancl lethargy Physical exam derecrs splenomegaly Blood work dis_

Desr

ro) accr) lcholine

(B) Block acetylcholine receprors

(C) Inhibit acetylcholinesrerase (D) Inhrbit acerylcholine sl.r-rrhesis

(E) Stimulate acetylcholine release inro rhe s),.napse

t5
You see a patient on an inltial visit and are struck by the blursh coloring of the skin and mucous membranes. you ask the parienr abour thls and you are told thar it is a bloocl problem that rhe parienr has had for his or her entire life. The patient's father had a similar condition, but not the
o1

mother. This conditron could result from w.hich one the followrng changes within the eryfirocyte?

increase of 2,3-bisphosphogiycerate in the erythrocyte (B) An E to V mutation at posrtion 6 of the B_chain of hemoglobin

(A)

An

(C) Increased oxldation of heme iron to the +3 (D) Enhanced oxTgen binding to hemoglobrn (E) A mutated hemoglobrn whjch no lor,rge.
the Bohr effect

state

exhibited

t6

\lany drugs funcrion by acting as inhibitors of particu_ lar enzyme reactions. If an enzl.meb i, li units/
%..,

Lt *:.:i:: l
f,

Hemoglobin

Glucose-6-phosphate dehydro genase C- Iron transport into the erythrocyte tD) Spectrin
(E) Methemoglobin reductase

tE

when a drop of rhe inhibitor flew inro his eye. This resulted in a pin-point pupil in that eye that was non_ reactive and unresponsive to atropine. He eventually (over a period of weeks) ...o,r.."d from this incidenl.
following? (A) Retraining of the ciliary muscles (B) Regrowth of neurons which were damaged by the
The reason for the long recovery period is which of the

Taboratory worker was working with a potent organo_ phosphorus inhibitor of acetylcf,olinesterase in the 1ab

jerking body movemenrs, an unsteady gait, personality changes, and chewing and swallowing difficulty, which has led to a gradual ,i.igt t loss. The patient's father had similar sl,riptoms before his death ar rhe age of 45. Cellular analysis indicated precipitated proteins in the nucleus. This disease has, at its origins, which biochemical problem? (A) An exonic delerion (B) A polygluramine rracr in an exon of the defective
gene

t patient has midlife onser of the following symproms: abnormal, involuntary

(C) A nonsense mutarion leading to the productlon of

truncated pro{ein

(D)

A splicing murarion, leading ro rhe insertion of

intronic sequences into the mature protein
protein producrion
of an unsrable mRNA, ieadlng to reduced

inhibitor

(E) Producrion

(C) Reslnthesis of the inhibited enzqe (D) Induction of enz),rnes which take the place of the inhibited erzyrr,e (E) Induction of proteases to reactivate the inhibited
CrIZ),rI].C

Protein Structure and

Function 13

{NSWERS
I
The answel is B: A noncompetitive inhibitor. Analysis ol the data indicates that in the presence of the inhibitor, the K, of the et:rzqe is the same as in the absence ol the inhrbitor, but th. V,.o. is significantly reduced (the extrapolated lines intersect on the x-axrs). These char-

with water and to ailow wa[er to maximize its entropr, It is not related to disulfide bond formation (cysrerne is not a hydrophobic residue) nor to hydrogen bond
formation of the side chains (hydrophobic side chains do not participate in hydrogen bonding). The clustenng
of hydrophoblc side chains may increase van der Waals interactions (.1ust by placing these residues in close proximity), but it will not necessarily minimize them. Sterlc hindrance between slde chains occurs as a protein folds, as the negative van der Waals interactions will prevent side chains from interfering wlth each other and the overall proteln stftlcture. The poly,rnerlzation of sickleceil hemoglobin molecules is due to hydrophobic interaciions between adjacent deoxygenated HbS molecules.

acteristics are the hallmark of noncompetltiye 1nh1bition, the inhibitor binds to a slte disrinct from the substrate binding site and aiters the proteins conformation such that activity is reduced, but not substrate binding. A competltiYe rnhlbitor would demonstrate an lncreased

Activation of the en4.me would either decrease the K or increase the V."., or both. Uncompetitive inhibitors
are very rare in pharmacology Such an inhibitor alters both the K and V-^, such that paralle1 lines are seen on double-reclprocal plots. The baslc concep[ behind rhis question is critical for an understanding ol hou- drugs alter enz).me activlties (the basis for pharmacolog}--). The answer is C: A very polar

\,,

but an unaltered V,"". (line intersection on the

_y-axrs).

The answer is D: An increase in hydrophobic interac-

tions between deoxyhemoglobin molecules. The boy is suflering from sickle cell anemia, rvhrch is due to a substitr-rtion o[ r-aline lor glutamate at position 6 of the B-charn This change lrom a negarively charged amrno acld srde chain \gluramate) to a hvdrophobic
side chain ir aline) aLLorvs deol-genate d hemoglobin to polyr-Lenze and form long rods u'rthin ihe red blood ce11. Deoxr.genated hemoglobrn has a hr-drophobrc patch on its surface (created b1.A70 FB5, and LSB), which the valine in position 6 on another hemoglobin chain can associate with via hydrophobrc interactions (this does not occur ln normal hemoglobin as there is a charged glutamate residue at this position, which will not lnteract with a surface hydrophobic patch-see the figure below). The bindlng of hemoglobin moiecules to

environment. The normal

pK, for a histidine side chain is 6.0, meaning rhat ar pH 6.0, 50o/o of the histidine side chains are protonated and 50% deprotonated. For the pK, to be raised to 8.2, there must be an envlronment which stabrhzes the protonated form of the side chain (because no\\. one has to reach a pH of 8.2 before 50% of the histidine side chalns have Iost their proton). A polar environment would stabilize histldine holding on to its proton, as compared to a hydrophobic enylronment, which would promote side chaln deprotonation at a low pH. The core of globular proteins is usually composed of hydrophobic amino acids (such as phenylalanine, valine, and leucrne), and in that environment, one would expect the pK" of the histidine side chaln to be reduced. Surface-assoclated domarns usually interact with water and are not where actlve sites are often lound (rt is too dlfficult to control the envlronment of the active site if water can freely enter the site). At a surface-associated domain, one would not expect much change in the histidine side chain pK. Many enzymes catalyze reactions based on the ability of amino acid side chains to accept or donate protons, which will be a function of the pK of the dissociable proton on the amino acid side chaln.

Strand no'1

fhe answer is B: Inrreasing the entropy of water. The

tendency for hydrophoblc slde chains to cluster ls driven by the entropy of water. Water will form a cage around hydrophobic molecules, which requires a decrease in water entropy The decrease ln entropy will be minimized, however, if water only has to form one large cage around a cluster of hydrophoblc molecules, rather than a
Iarge number of smal1 cages around separate hydropho-

Axial contacl

63A

Gtu

tZt

2Fz Thr 4

2a,

bic molecules. Thus, the drir,rng force for the hydrophobic side chains to cluster is to mlnlmize their interactions

Hydrophobic interacLions involved rn deoxyhemoglobrn S forming

long pol1.mers.

r4

Chapter 2

each other results in the polymerization. Oxygenated hemoglobrn does not present a hydrophoblc surface to other hemoglobrn molecules, so polymerization is much less hkely in the oxygenated state. The poiymerization is not caused by a loss ol quaternary structure, an increase in oxygen binding (whlch would actually reduce sickling), a gain of ionic interactions, or the loss ol any u-helical structure in the fina1 conformation of the protein.

The answer is D: A misfolded lorm of a normal protein.

The pathologist is showlng early clinlcal signs of

Creutzfeldt-Jakob disease, caused by a misfolded prion protein, leadlng to protein aggregates in the brain The lnitial seed for the aggregation was obtained lrom a cadaver that the pathologist was lvorking on Prions can exist in two states, the normal, nonaggregaled form and an aiternatrve conformation that rs prone lo aggregation (see differences in structure be1ow, where PrP' is the normai conformation, and PrP" is the abnormal structure). Once the allernative form reaches a cntical concentration, aggregation ensues and shifts the equrlibrium between the normal and abnormal lorms lo produce more abnormal form, feeding the aggregatron The prion is not a truncaled neuronal protein (its pnmary structure can be the same in both forms ol the protern), nor is it a truncated extracellular protein Thrs disorder is not due to alterations in hemoglobin or fibri111n. Thls patient will probably die within 1 year. There is no current treatment for the disease. As the disease progresses, he wiil probably develop blindness, involuntary movements, and severe deterioration ol mental function.

mutations in flbril1in, an extracellular protein. Fibrillin helps to form, along with other proteins, microfi.bri1s, which are present in elastic fibers (containlng primarily elastin), which help to give r.arious lissues thelr elastic properties. The exact mechanism whereby mutalions in fibri11in iead to the symptoms of Marfan s;,r-rdrome has yet to be established. MuLations in any ol the other proteins listed do not glve rise to Marfan's (although co1lagen delects give rise to osteogenesis imperfecta, and dystrophin mu[ations give rise to various lorms of muscular dystrophl,, depending on the type of mutation). Marfans is an autosomal dominant dlsorder of connecti\.e tlssue (not collagen). It is caused by mutations in the FBN 1 gene (located on chromosome 15), which encodes fibrlllln-1, a glycoprotein. The picture rs of a dislocated
lens, a classical finding in patients with Marfans.
The answer is D: Decreased resorption of collagen. The patient has a form of osteogenesis imperfecta, which is due to a mutation rn collagen, generating brittle bones.

Mild trauma is sufficient to break the bones. Blsphosphonates decrease bone resorption by the osteoclasts, thereby strengthenlng the bone, even with the defective collagen molecule. Brsphosphonates do not affect the s;,T"rthesis of
collagen or fibn1lin. The answer is A: Antibody Iight chains. The patlent ls exhibiting the symptoms of primary amylordosis, which is a protein folding disease in which immunoglobulin light chains are improperly processed and cannot be degraded. These proteins then form fibrils in tissues,

which are insoluble. This disrupts the normal function of the tissue, and many tissues can accumulale these f,bri1s. Primary amyloidosis does not occur wrth abnormal deposlts of co11agen, fibrillin, albumin, or serum
transaminases.

of 2,3'bisphospho' glycerate. 2,3-bisphosphoglycerate (2,3-BPG) will bind to and stabilize the deoxygenated form of hemoThe answer is C: lncreased synthesis

PrPc

PrP""

Note rhe difference in struclure between PrP' (normal; three major helices and two minor B-sheets) and PrP" (abnormal; four major B-sheets and two major helices). The abnormal form is much more prone to aggregate, due to the alterations in tertiary structure.

The answer

slrnptoms

is B: Fibrillin. The boy is showing the ol Marfan sytrdrome, which is caused by

giobin. Thus, if 2,3-BPG ievels are increased, the hrndrng ol this molecule wt11 aid in removing oxygen lrom hemoglobin in the tissues (where the concentration of oxygen rs 1ow) and therefore increase oxygen dehvery to the tissues. In the 1ungs, where the oxygen concentration is high, the high levels ol oxygen can overcome the effects of 2,3-BPG and brnd to hemoglobin. Lactic acid 1eve1s do not directly affect oxygen binding (and lactate does not accumulate ln the red ce11), although changes in proton concentration (pH) can Decreased pH rnil1 reduce oxygen bindrng to hemoglobln due to the Bohr effect. Bilirubln degradation, even though it does produce CO, does not effect oxlrgen binding to hemoglobin.

Protein Structure ;rncl F-.:-,

10 The answer is C: Hydroxylation of proline residues in collagen. Limes provided r.itamln C, r.l,hich is a required -.l ector lor prolyl hydroxl lase, the enzl.me r,vhich -'. Jrorl-laies proline residues in co11agen. L).sine cross-

1;

shown

Other r.ariants of the helix are 3/ltl anr - belor'r,-. rn a linear fashion. are rh. :bonds formed in aLI three type s of hehces

-ris in collagen do not require vitamin C (although ' srne hydroxl.lation, for the purpose of glycosylation, :,--ls). \ritamin C does not affect calcium mobihzation

3.Cri 0

::at is vitamin D) and fibri11in is not the problem

.:: ritamin C deficiency Glycine residues rn coliagen
--',nnot be conr,'erted to proline r,vithin the pol,vpeptlde.

3.6/13
lrv

I1

The answer is D: lnability to carboxylate glutamic acid side chains. Cystic fibrosis patients har.e a thickening

--

ilIIltrIilIIilIIilIIIII .C_N_C_C- N-C_C_N_C-C- N- C_C-N_C_ C_ Nttttt r11 R2 R3 R, R. l+

The answer is B: BIock acetylcholine rereptors. The palient has myasthenia gra\is, in u,hich she generates antibodies against the acetylcholine receptor. TieatmerLt r,vith edrophonrr:m chloride 1eac1s to a transirnr incrcasc in ace tvlcholine levels (Lhrough the tempoi-eq-inrctrvrLion ol acetylcholinesterase) such that acetl.lcholine can brnd to receptors (r.ia compeLrtion with the antrbodies). Normai levels of acetl,lcholine are too low lor such competltlon to be successlul This disorder does not generate antibodies lvhich lead to acetylcholine destruction, lnhi,

OHHOHHOHHOHHOHHOH

tl'Le

pancreatic duct, leading Lo nutrient malabsorp,

, ,1n. as pancrealic enzymes have

intestinal lumen Liprd malabsorptron sl,ndromes .lquently lead to de{rciencies in fat-soluble r-itamin -.:take (r.rtamins E, D, K, and A). Vitamin K is required -.r the carboxylation of glutamic acid side chains on :iood clotting proteins. This provides a means for -:-rese proteins to chelate calcium, and to bind to plate:t surlaces. In the absence of gamma-carbory,latron ol _.iutamate, the clotting complexes cannot form, and :. clotting drsorder is observed. Vitamin C is requirecl --.r proltne h).droxylation, and as vitarnrn C is a u.ater-,.1ub1e vitamin, lipid malabsorption does not allect
'..ttamin C uptake. Transaminations require r-rt.rmin B,, :.:.rother rvater-solub1e r.itamln. Dolichol can be synthe.i,ed in the bodl: so its absorption is not an issue uncier :hese conditlons. Endogenous fatty acids wr11 provide

.rt

difficulty reaching

bition of acetylchohllester-ase , inhibition of acetylcholine

s),nthesis, or release of acetylcholine at the synapse.

l5

The answer is

(: lncreased oxidation of heme iron to the +3

methemoglobinemia.

.nergy lor protein

'ra

s1,r-rthesis

rn rndrviduals with lipid

state. The patient is exhrbiting

labsorpt ion problems.

t2

The answer is E: lane 3 represents an individual with sickle cell disease. The mutation in sickle cel1 drs-

:ase ls a r.aline for glutamate substitution at posltton c ol the B-chain. This substitution removes a negati\-e :harge from the B-chain such thaL when the B-chain is n.iigrated through an electric lield it will not travel as tar towards the posltive pole as does the nonmutated proteln. Thus, in the gel shorvn in the question, band X i.presenLs the hemoglobln S B-charn (since it does not nrgrate as far towards the positlve pole), and band Y :epresents the nonmutated protein. The pattern shou.n rn lane 1 is that of a carrler of HbS (one normal p-chain and one mutated p-chain) The pattern shown in lane 2 represents a person who does not carry a mutant a11ele .tu-o normal alleles). Lane 3 represents someone with the disease (two mutant genes).

in rvhrch an rncreased pe rcentage of his hemoglobrn has the iron in the +3 oxrdatlon state (normal ls +2), which is a lorm that cannot bind oxy,gen. Thls condition ean arise by a variety ol mutations within hemoglobrn rvhrch lead to destabilization ol the iron in the heme nng. The red blood cell contains methemoglobin reductase, u.hich r,rrll reduce the iron back to the +2 state (using NADPH as rhe electron donor), and mutations within the reductase can also lead to this condltlon. An acquired form of methemoglobinemia can be caused by exposure to oxidizing drugs or toxins (aniline dyes, nttrates, niirites, and iido, caine) which exceed the reduction capaciry of the red blood ce11s. Surprlsingly,, the malority of patients urth this sl.ndrome show no i11 eflects, other than the bluish discoioratlon of certain tissues. Excessive 2,3-bisp1-rosphoglycerate in the erythrocyte would lead to increaseci oxygen delivery to ihe tissues as 2,3-bisphosphogh cerate stabilizes the deoxygenated lorm ol hen-rogkr1,-1n (as would a mutant l-remoglobin with an enhanced rrbr'ity to bind 2,3-BPG). The E to V mutation at pLrsitlLrn r the B-chain ol hemoglobrn leads to sickle ce 1l dLsease

t3

The answer is C: B and In a t1.pica1 cr-helix, there are 13 atoms between hydrogen bonds (formed between the

E.

:arbonyl oxygen of one amino acid and the amide nitro:en of the amlno acid four residues up in the chain) Thls is referred to as a 3.6/13 helix (3.6 amino acids :er turn, with 13 atoms between hydrogen bonds).

16

The answer is B: 4 units/min/mg protein.

Thi-. rs sri-, .

usrng the Michaelis-Menten equation. L(,,2S11 Under inhlbiied conditions \/.

,

r6

Chapter 2

mg protein, the K* is l.25pM, and the substrate concentration is 2.50pM. Plugging these numbers into the
equation leads to a value of v of 4 units/minimg protein.

l7

The answer is D: A defect in the structure of the sarco' Iemma. The boy has Duchenne muscular dystrophy,

which is due to mutatlons in the proteln dystrophin,

found in the muscle sarcolemma (plasma membrane). The lack of dystrophin aiters the permeability properties of the plasma membrane, eventually leading to ce11 death. The disorder is not found in mitochondria,
the liver, or in the B-chain of hemoglobin. This disease also does not alter gene transcription. As the muscles weaken, their function is compromised, leading to the complications of this form of muscular dystrophy.

t8 fhe answer is D: Spectrin. The child is exhibiting

the symptoms of hereditary spherocytosis, a defect in
A
Band 3 protein

spectrin in the erythrocyte membrane. This membrane problem ieads to an abnormal shape of the red blood ce1i, such that the spleen removes them from circulation (hence, the large spleen), Ieading to an anemia due to a reduction of red blood cells in circulation. This defect is not due to a loss of hemoglobln or glucose-6-phosphate dehydrogenase (a lack of glucose-6-phosphate dehydrogenase will lead to red cell damage and cell fragments on peripheral smear under oxidi.zing conditions, conditj.ons not observed with this patient). A lack of iron in the erythrocyte can lead to an anemia (due to insufficient oxygen binding to hemoglobin and reduced o*ygen delivery to the tissues), but it would not lead to an altered ce11 shape. A loss of methemoglobin reductase wouid lead to increased levels of methemoglobin, which cannot bind oxygen, but would also not lead to a cell shape change. The placement of spectrin in the red cell membrane is shown in the figure.

19

The answer is (: Resynthesis of the inhibited enzyme. Once acetylchollnesterase has been covalently modified by an inhlbitor, it cannot be reactivated. The only way to

regain thls activity is by new slrlthesis of acetylcholinesterase, which would not have the covalent modification found in the inhibited enz]Tne. Since acetylcholine is released at nerve muscle junctions, once new acetylcholinesterase has been slmthesized the released acetylcholine can be cleaved in order to allow relaxation of
the muscle.

ci-spectrin

P-SPectrin

20

fhe answer is B: A polyglutamine tract in an exon of the defective gene. The patient ls suffering from Huntington disease, which is transmitted in an autosomal dominant pattern in which a triplet repeat is expanded within the Huntington dlsease gene. This triplet repeat codes for a polyglutamine lract in the maiure protein, which leads to its eventual failure and disease sl, nptoms. Huntingtons is not caused by an exonic deletlon or a nonsense mutation. Splicing ls normal for the gene, and the mature mRNA is stable.

Chapter 3

ffiNA

Sesesrffiff*=

RepErym*EBF

affid R*Fary
Tlrc- questians

.i

,it*tations could rssult, hannitag tht slsedes, sbilitt, .: stttt,iye in future gtmwretia*.s. As srlcla, repaifitzg ,:i1.ors_dwing replkatian *wd regtsiting e*"at"s thil. : ccur before replicatian (*s iwdttcidbl, eiz,iyonmcmt*! g en ts) a r e cricial o r the sp e ci e s, * i g- t r. ttzr s *rv, jvra i. f Tlis clupter ljft:setts quistians catlt;rrning a wids ofiety of topics relating to this thewe.
I

ibility to think through qitsti*ts relritlitg t* D,1A. is DN,A is the human genttir tttcttst-i_{tl, * *u-ist be 'tplicatedfaitWy; othet **ise . y*ttntiai rltletsrlous

in

ffuis chatrst* exr*szit,,{

a stLLa{iLt.s

Tl-re

number ol compiementaiion groups represented by

these parienrs is

l-hich ol the follou,rng?

t-{r 1 rB) 2 (c) 3

(D) 4 (E) 5

Analysis of a celi line that rapidly transforms into a rumor cell line demonstrated an increased mutation rate r,r,ithin

QUESTIONS
Select the single best answer.

tion in the DNA polymerase enzyrne that sprthesizes the

the cel1. Further analysis indicated that there was a mura_

in which of rhe foliowing activities of this DNA

merase?

leading srrand. This inactivating mutatron is 1ike1y to be

poly_

A ciinic lvas studying patients with xeroderma pigmen_ tosum and ran experiments to determine how many
different complementarion groups were represented in their patient sample. Fibroblast cell lines were creared lrom five different parients and fused with each other (al1
possible fusions were examined, as shown in Table 3_l).

(A) 5'- l' exonur.lease actrvity (St l'-5' exonuclease activity

(C) Phosphodiesrer bond making capability (D) Uracil-DNA glycosylase actrvity

(E)

Ligase acrivity

The resultant heterodikaryons were then examined for therr resistance to UV light, as indicated below (a ,,+,, indicates resistance to UV damage, while a ,,-,,indicates sensitivlty to UV damage). Table 3-1.
Cell

Number1234S

DNA damage, but only in the absence of folic

[ollor,i ing?

13-year,o1d exhibited developmental de1ay, learning disabilities, mood swings. and ar times, autrsric behav_ ior when he was younger. His current ph;.sical exam shows a long face, large ears, and large, fleshy hands. His fingers exhibir hyperexrensible loinrs. Eraminarion of fibroblasts cultured from the boy shou,ed abnormal
acrd.

This disorder has, at the genetic leve1, which one of the

(A) A single missen>e mur.rIion (B) A large deletion

(C) An exrended triplet nucleotlde repeat (D) A nonsense mutatlon

(E) L7

Gene inaciivation r{a methr4ation

ffi,8

Chapter 3 pediatrician,s

ii#:; An S-month-old child is brought ro rhe

olfice due to excessive sensitivity to the sun. Skin areas exposed to the sun for only a brief period of time were reddened with scaling Irregular dark spots have also appeared. The pediatricran suspects a generic disorder in which of the following processes? (A) DNA replicarion (B) Tianscription (p Base excision repair @)Nucleor ide excision repair (E) Tianslation

:,:E

deamination can lead to mutations if not repaired. Which deaminarion indicated below wouid iead to a mutation in a resulting protein if not repaired?

Spontaneous deamlnation of certain bases in DNA occurs at a constant rate under all condlttons. Such

#
NHz

QHzOH

(B) ctou
(C) GtoA (E)

(A) Ttou

H-r.r\r

(D)Atoc Utoc

"L.)
I

a,9

sees an obsretrician due to difflculties of the woman keeping a pregnancy to terrn. She has had three miscarriages over the past 6 years, and the couple is searchlng for an answer. Karyot1.pe analysis of the woman gave the result of 45,XXder(L4;21). A likely potential cause of the miscarriages may be which of the following? (A) Imbalance of DNA in pollploid conceprions (B) Imbalance of DNA in euploid conceprions_ (C) Triple X concepr ions

A couple

geal candidiasis. She had a hlstory of drug abuse and needle sharing. Blood analysis indicated a CD4 l.wr'_ phocyte count of less than 200. Whlch of the foliowing compounds would be a drug of choice for this patienti

(D) Zero X conceprions (E) lriro-, 2[ conceptions

?.,:f"E A32-year-old woman exhibited a

#::, ,n"high mutation rare of the human immunodeficiency (HIV)

virus is due in parr ro a properry of which of following host cell en4.,mes?

thl

high fever, malaise, generalized lymphadenopathy, weight loss, and esopha-

(A) DNA polymerase ,@ nNa poil,rnerase (C) DNA primase (D) Telomerase (E) DNA iigase
i,:jjlFj Consider the DNA replication fork shown beiow. DNA ' - " "' ligase will be required to finish qmthesis at whlch labeled points on the figure?

B
//

NH2

N)

NN

/r-Nt\

DNA Structure, Replication, and Repair

l9

(A) A and

(B) C and D
(C) A and C

(A) Brse excision repair (B) DNA replication

(C) Tianscription-coupled DNA repair (D) Proolreading by DNA polymerase

(D) D and

(E)

B and C

(E)

Seahng nicks in DNA

The sequence of part of a DNA strand is the follou,ing: -ATTCGATTGCCCACGT-. When thls strand is used as a template for DNA rytthesis, the product will be u.hich one of the following?

t4 A woman visits her physician due to fever and pain upon urination. Urinary analysis shows bacteria,
leukocytes, and leukocyte esterase in the urine, and the physician places the woman on a quinolone antibiotic

(A) TAAGCTAACGGGTGCA (B) UAA.GCUA-{CGGGUGCA (C) ACGUGGGCAAUCGA-{U (D) ACGTGGGCA\TCGA\T (E) TGCACCCGTTAGCTTA

(ciprofloxacln) The mammalian counterpart

to

the

bacterlal enzyme inhiblted by thrs drug is rvhich of the follou'ing? (A) DNA polymerase cx

tl

You have been following a newborn lvho first presented

with hypotonia and trouble sucking. Special leeding techniques were required for the child io gain nourishment. As the chl1d aged, there appeared to be der-e1opmental de1a1', and the child then gained a great inleresr in eating, and rapidly became obese Der-elopmenral delay was still er'rdent, as was hypotonia. A karyotl'pe analysis of this patient would indicate lvhlch of the fo1lowrng?

(B) Topoisomerase (C) Llgase (D) Pnn'rase (El Helicase t5 \\ hich

ans\\

be

Lori besr predicts the effect of the lo1-

lourng drug on the pathu-ar-s inchcatecl?

(A) A monosomy (B) A trisomy (C) A duplication (D) A chromosomal inversion (E) A deletion

CH,_OH

t2

You see a 2-year-old child of Ashkenazi Jewish descent who is very small lor her age. The patient exhibrts a long, narrow face, sma1l lower jaw, and prominent eyes and ears. The child is very sensrtirre to berng outdoors

in the sun, often burning

easlly,

with butterfly-shaped
DNA
RNA

patches of redness on her skin. Upon testing, the child rs also slightly developmentally delayed. The delectlve protein in this chl1d is which of the following?

Protein

Synthesis
(A)
(B)
ln

Synthesis
lnhibit
No effect

Synthesis
No effect No elfect No effect No effect Inhlbit

(A) DNA polymerase (B) DNA ligase (C) RNA polymerase (D) DNA helicase (E) Reverse transcriptase
13.. Concerned parents are referred to a speciaity chnic by their family physlcian due to abnormalities in their 1S-month-old childs development. The child displays delayed psychomotor der.elopment, and is mentally
retarded. The child is photosensitirre, and also appears to be aging prematurely, wlth a stooped posture and sunken eyes. The altered process in thls autosomal recessir.e disorder is which of the followrng?

hibrt

lnhibit No elfect No effect No eflect

(e)
(D)
(E)

lnhibit No effecl No effect

tG: A new patient

visits your practice due to hrs concern of developing colon cancer. A large number of relatrves have had premature (1ess than the age of 45) colon cancer, and all cases were nght-slded, wrth the only visible polyps being found on that side. The molecular basis for this form of colon cancer is which of the foilowing?

(6) A defect in DNA mlsmatch repair

1B) A delect in base excision repair (C) A defect ln the Wnt signaling pathway (D) A defect in repalring double-strand DNA breaks (E) A defect ln telomerase

cells detects the presence of the following karyot1pe. The molecular basis of this dj.sease is which of the following? (A) Loss of an essential tumor suppressor activity (B) lncreased rate of DNA mutation due to loss of DNA repair enzymes

(C) Creation of
cells

a fusion protein not normally found

in

t1

Over 50olo of human tumors have developed an inactivating mutation in p53 activity The lack of this actiuty coniributes to tumor cel1 growth via which one of the

(D)

Loss of a critical tlrosine kinase activity

(E) Gain of a critical ser/thr kinase activity

20 A scientist is replicating human DNA in a test tube and has added intact DNA, the replisome complex, and the four deoxyribonucleoside triphosphates. To the surprise of the scientist, there was no DNA q,nthesized, as determined by the incorporation of radio-1abe1ed precursors into acid-precipitable material. The scientist's failure to slnthesize DNA is most likely due to a lack of which of the following in his reactlon mj.xture?

loliowing mechanisms? (A) Loss of Wnt signaling (B) Increase in DNA mutation

rates

1C) Activation of MAP kinases (D) Increase ln apoptotic events (E) increase in transcription-coupled DNA repair

t8

The isolation of nascen[ Okazaki fragments during DNA

replication 1ed to the surprlsing discovery of uracil in the fragment. The uracil is present due to which of the
following? 14) Deamination ol cytosine (B) Chemical modification of thy'rnine (C) An error in DNA poll.merase tD) Failure ol mismaich repair @) fhe need for a primer
19
You have a patient with an elevated white blood cell count

(A)

Reverse transcriptase

(E)

Ribonucleoside triphosphates

(C) Templates (D) Dideoryrrucleoside triphosphates

(E) Sigma factor

and a feeling of malaise. Molecular analysis of the whlte

sr
$

**

$q *
,.-*

22f
Phl

DNA Structure, Replication, and Repa,r

2l

{NSWERS
t
The ansruer is C:

3.

Cel1

lines complement each other

The answer is C: An extended triplet nucleotide repeat. The boy rs displaying the symptoms of fragile X sr:-

il

their mutations are in different genes. For the pur-

pose of this questlon, 1et us assume there are three genes involved, lettered x, y, and z. Cell line I is def,cient rn

but since it can complement every other ce11 line, ce1l lines 2 through 5 cannot be deficient in gene x. When fused, the other cel1 lines (2 through 5) produce normal x protein, whrch complements the deficiency rn cell line l. Similarly, cel1 iine I produces normal copres of the genes that are deficient ln celi lines 2 through 5. Thrs indicates that there are at least two complementation groups available. Cell iine 2 complements cel1 lines l, 4, and 5, but not 3. Thus, the mutation in cell line 2 (ca11 it gene y) is also present in cell line 3 (slnce the two cel1 lines cannot complement each other), but not rn cell lines 4 and 5. Thus, at this point, cell line 1 is deficient in gene x, and cell lines 2 and 3 are defrcient in gene y. Cell line 4 complements cel1 lines 1, 2, and 3, but not 5. Thus, cel1 1lnes 4 and 5 have similar mutations, but in a gene distinct from genes x and y Thus, cel1 lines 4 and 5 can be deficient in gene z. The cell lines are thus 1 (x-), 2 and 3 (y ) and 4 and 5 (z ). So, as an example, when cell line I is fused with cell line 2, cel1 line I is x-y*, and cel1 line 2 is x*y , so the fused ce11 (x y*/x*/y-) produces both normal x and y
gene x,

drome. Fragiie X contains a triplet nucleotide repe;.', (CGG) on the X chromosome in the 5' untranslat.d region of the FMR1 gene. The triplet repeat expansion ieads to no expression of the FMR1 gene, lvhich produces a protein required for brain development. Its function appears to be that of an mRNA shuttle. moving mRNA from the nucleus to appropriare sites in the cytoplasm for translation to occur. Depending on rhe level of expression of FMRi (which is dependent on the number of repeats), the symptoms can vary

from mild to severe. Less than 1% of cases of fragile X are due to missense or nonsense mutations; the vast majorlty are due to the expansion of the triplet repeat
at the 5r end of the gene. Gene inactivation by methylation, or deletion, are not causes o[ fragile X syndrome. The sprdrome was called fragile X because the X chromosome that carries the repeat expansion is subject to

DNA strand break under certain condltions (such as

lack of folic acid), which does not occur with normal X chromosomes. The area contarning the repeat alters the stalning pattern of the X chromosome, allowing this to be seen in a karyotype (as seen below). Fragile X is the most common inheritable cause of mental retardation. Males are more severely affected. In early chlldhood, developmental delay, speech and language problems, and autisticlike behar.ior are notlceable. After puberty,

The answer is B: 3'-5'exonuclease activity. DNA polymerase rarely makes mistakes when inserting bases into

the classic physical signs develop (large

prominent jaw).

testicles,

Iong-thin face, mental retardation, large ears, and

a newly s1,r-ithesized strand and base-pairing with the template strand. However, mistakes do occur at a frequency of about one in a million bases sy, rthesized, but
DNA
po11n-Lerase has an

error checking capability which

base before proceeding with the next base insertion. This is due to the 3'-5' exonuclease activity of DNA poll.merase by which, prior to adding the next nucleotide to the growing DNA chaln, the base put into place in the previous step is examined for correct base-pairing propertles. ]f it is incorrect, the enabies

it to remove the mispaired

en4rrne goes "backwards" and removes the incorrect

base, then replaces it with the correct base. The 5'-3' exonuclease activity of DNA poll.rnerase moves ahead, and is used to remove RNA primers from newly sy'nthe-

#
ff
f ratxl Y
Picture

w
ffi

sized DNA.

If the en4.me could no ionger

s)-nthesize

phosphodiester bonds (the primary responsibllity of the en4.,rne), DNA slnthesis would halt. A loss of uracil-DNA
glycosylase activity is not a property of DNA po11.merase, but that of a separate enzqe system which repairs spontaneous deamination of cytosine bases to uracil within DNA strands. If these were left intact, mutations wouid increase in DNA. A loss of ligase actir'rty would lead to unstable DNA, as the Okazaki fragments would not be able to be sealed together to form one continuous prece ol DNA, and this would most 1ike1y lead to cel1 death, not an increased mutation rate.

# ffir xy

ol a fragile X

chromosome and normal X and Y chromo-

somes. Note the end of the long arm (q), and the diflerences between

the two chromosomes.

ffi

ilF:ii:1!:::it

22

Chapter 3
The answer is D: llucleotide excision repair. The child rs suffenng from a form of xeroderma pigmentosum, a disorder rn which th)..rnine dimers (created by exposure

5t

W light) cannot be appropriately repalred in DNA. Nucleotide excision repair enzymes recognize bulky dlstortions in the helix, whereas base excision repai.r recogto
nizes only specific lesions of a smali, singie, damaged base.

The mechanism whereby th)'T nine dimers are removed from DNA is nucleotide excisron repair in which entlre nucieotides are removed from the damaged DNA. In base excision repair, only a single base is removed, the sugar phosphate backbone is initraliy left intact (see the figure below for comparisons between these two systems for repairing DNA). This disorder is not due to alterattons in transcription (sprthesizing RNA from DNA), DNA replication, or translation (syrthesizing protelns from mRNA). Another example of a disease resulting from a defect in nucleotlde excision repair is Cockalne sprdrome. Neurological dlseases (such as Alzhelmer's) may also have a deficiency in nucleotjde excision repair. Normal DNA

The answer is B: C to U. Cytosine spontaneously deaminates to form uracil while ln DNA. Thls error is repaired by the uracil-DNA glycosylase system, which recognizes this abnormal base in DNA and initiates the process of base excision repair to correct the mistake. Neither thymine nor uracll contains an amino group to deaminate (thus, answers A and E are incorrect). When adenine deaminates, the base hypoxanthine is formed (lnoslne as part of a nucleoside), and guanine deamination will lead to xanthine production. The deaminatron of cytosine and conversion to uridine is
shown be1ow.

NHz NHs

"t)
I

,,

:$
I

-1r::-r1::
.i:;'&:1:.".:]:!"l'::J...*

The deamrnation of cytldine to uridlne (C to U

wthin a DNA

strand).

r'
TE:::::
i--!-:!-+-:l;
I

Zurun\ tobases

The answer is B: lmbalance

of DllA in euploid

concep'

tions.

6 t'{
I

glvcosYtase

v
1j1:.-"":]I=
B
s.4,f5-:+4-'

N
U

c
I

:
: Y;
c
i

t
-2!.,&-!-:-+

incisionendonuclease

*T&*
/-1_-e''-f M 1-_:-*'!

t
i

The woman has a Robertsonian translocation between chromosomes l'1 and 21 (the two chromosomes are fused together at their stalks; see the flgure on page 23). When she creates her eggs, there is an imbalance in the amount of DNA representing chromosomes 1'l and 2I in the eggs, such that fertllization of the eggs will lead to either monosomy or lrisomy with these chromosomes, most of which are lncompatible with life. The figure below indicates these potential outcomes. Polyploid outcomes would be three or more times the normal number of chromosomes, which does

:
e

x
c
i

not occur here; and the Robertsonian translocation will not affect the distribution of the X chromosome. Tiisomy 21 witl lead to a hve birth, Down s1'ridrome,
although there is sti11 a risk of mlscarriage with trisomy 21 conceptions. The risk is lower, however, than an imbalance of DNA brought about by the segregation of the chromosomes containing the Robertsonian translocation. Euploid cel1s have a number of chromosomes which are exact multiples of the haploids (in humans haploid is 23, diploid is 46, and polyploid is 69 or 92
chromosomes).

;
e
p
I

S
i

o
n

a
i

I DNA polymerase

:xY/{r:, i;;x:(;;*'
I

L
p

Nick

a
i

IDNA liqase

Tre

;xJ-Xxx*i
Repaired (normal) DNA A comparison of nucleotide excrston repair and base excision reparr

DNA Structure. Replicarictn. and i.;:-

)3

Balanced

14

*& fr&:'. i4
U
4;2'1

21

&.

"'

21

; xsse. l& * &;.' ,-$

13

45.XX,der(1

){q 1 0;q 1 0)

45.XX,de

3;14)(q1 0;q1 0)

Unbalance.i , .&ii 21 t4Y;X"14*

*#

13#
&

21
46,Xy,der(14;21)(q
t

21

'" 6* e& ..# **

13 13
14

0),+2.1 Down's syndrome

(46,xX,+ 13,der(13;.14)(q t 0;10) patau's syndr6me

\i Affi AA-*/=\----.---* x v Q'e ur,s* 2, $}fl$-'?::i'#jf"?'

B'sater;1s6

rllrmar

{ -. \ ra.anozr
lMerosistconfrgJ.arion

\:;"."",,""*j::ll',."",,,,
r-''=-v
=*, mm

o.,rpe.mrrscen,r;ciusion

iffiH

.rrer 6: The most

: lt) (lefr) and der(13;14) .:tr (A,B). A meiosis I con-

-:.,:anlan transiocalions

important
are Possible gametes

$$
Normal

il ,j

I]/t )l u it il
u ii
ii

Il

fl
hl
EJ

-,. .r1 fbrmed in a carriet of -.21) is shol-n in panel

Carrier

dl HI]
trisomy
0 0

?iJ
14

lt
U

:lq

. :
r
.

tith

Zyqote
outcome
Observed proportions: Maternal carrier

t
I

Translocation trisomy 21

Monosomy
21

Translocation Monosomy
14

the six possible

products (D), of r.vhich :-:'te are er.er obsened. Fresiaristics are based on pre-

t
der t(14;21)
D

lDown's syndromel some miscarry

IDiesj

lMiscarriagel

lDies]

35% 387"

50% 60%

15%

0 0

0 0

----..qnosis results rn

carriers.

Palernal carrier

The answer is D. The woman is suffenng from AIDS, -..nd one class of drugs used to srop the spread ol the -.-lrus is the drdeoryr-rucleosrdes (the compound shown

dideoxyadenosine). The dideoxl,nu,leosides interfere r.vith DNA sy.nthesis after they are -..itn'ated to the triphosphate leve1 through purine sa1".,rge pathway enzyrnes. Since ihese compounds lack ,, 3'-hydroxyl group, once they are incorporated into -. grou,ing DNA strand, they cannot form a phospho..resler bond with the next nucleotide, and slmthesis :iLrps. Reverse transcriptase, an enz)-me carried by HIV

,n answer D is

rit

not found in eukaryotic cells, appears to have

-rgher alfinity for these drugs than does normal cellular )\A polymerase, so the agents har..e a greater ability -: preferentially stop r,rrus sl,rthesis and not cellular

DNA synthesis, although it does occur to a sma11 extent. Structure A is adenosine, a ribonucleoside which when actlvated may be used for primer synthesis in DNA replication, but not as part of the DNA structure. Strucrure B rs methotrexate, an agent which inhibits drhydrololare reductase and blocks the slnthesis of thymidine, therebr blocking DNA s1-nthesis. lt is used as a treatment for psoriasis and r,vas used, in the past, as a chemothera peutic agent. Structure C is deoxyadenosine, lvhich is a normal substrate for DNA polymerase after acrrr:tion. Structure E is 5-fluorouracil (5FU), an inhibttor ..i thymidylate sl,rrihase. 5FU blocks rhymidine srlrhc-s,s and stops overall DNA s1.r'rthesis. It is used for uen.rLttumors as an anticancer drug, but is not use d tor Hi
.

rnlections.

24

Chapter 3
The answer is B: RllA polymerase. During the life cycle

change that confers a growth advantage to this strain of

of the HiV the double-stranded DNA whlch was produced from the genomic RNA integrates randomly into the host chromosome (see the figure below) Ce11ular RNA poll,merase lhen transcribes the viral DNA to produce viral RNA, which is used in the translation of viral proteins, and as the genomic material for new virions' RNe pollT t erase lacks 3'-5' exonuciease activity, thus the enz),rne cannot correct any errors it may make while transcribing the viral DNA. The RNA produced, which carries errors in transcription, is then packaged into a new virus paftic1e, and this muta[ion may lead to a

virus. The lack of proofreading by RNA

po11'rnerase is

many mesnot usually a problem in eukaryotic gene, and lf 1olo of those sages are produced from a singie messages produce a mutated protein it will be compensated by the 99ok of the messages which produce a norma1 protein. In the viral case, however, the mRNA turns lnto the genomic material, which will lead to mutations in ail future descendants of that virus. This is why HIV is treated with multiple, different antivirals simultaneously, to destroy any virus whlch mutates to be resistant to the antiviral agents. DNA polltnerase has error-checking
ce11s, as

HIV life cycle

-ii HN bhds to theT celi.

.:t Vnal NA

is

releded

hio

iil Revelse lranscdPiase conveds viral RNA hto vilat DNA.

+ Vnal DNA j,

enters the T-

ce['s nu.leus md inseils itself hto th T-cell's DNA

The T cell begbs to make copies of the HV

l:r Protease (an eroYme) helps 0eare new virus pardcles.

:', The

new

w vljon (ttus
is released

padde)

ftom

&e T-celi.

Vir.l DNA

New

N virion (vhs particle)

HN protehs

The HIV life cycle

DNA Structure, Replicatlon, and Repair .apabiiiries, and will nor significantly increase the muta_ tion rare of the integrated viral DNA. DNA primase may make errors, but they are corrected when the RNa pnmer is removed from the DNA. Telomerase only works on the ends of chromosomes, and the viral DNA does nor usu_ a1lr' integrare at those positions. DNA ligase activity is not required for viral RNA production.
The answer is E: B and

25

C.

tron in which the replication fork is moving.

5'

lagging s-trand synthesis in these two replication forks \see the figure below). This means that the DNA rs s1,n_ thesized in the direction opposite to thar of the direc_
Because

The areas labeled B and C are

known as Okazaki fragmlnts. These Okazaki fragments need to be sealed together, which occurs with DNA ligase (after the RNA primers have been removed by a DNA polFnerase wrth a 5,_3, exonuclease activity). The vertical line refers to the origin of replication, and labels A and D are the leading ,t.*d, of DNA slrrrhesis, which, since syrrthesis is occurring in the direclion that the replication fork is moving, can be sy,nthesized
continuous piece of DNA.
as one

direction)

of this, the DNA must be sl,nthesized in shorr preces (as DNA polyrnerase can only sprthesize DNA in rhe 5' to 3' direction, reading the template in the 3, to j,

3,

to *
s'',t,.

5,
3,!: \ ,

3,

l0

o \RNA
Prlmers

-i

The answer is D: ACGTGGGCAATCGAAT. The product of

DNA replication wiil be complemenrary

'

't

't.. 1t-*1 First round

of synthesis (O)
3':..----1 5'

=.i
I

remplate, the product will be 3/_TA{GCTATCGGG} GCA-. When writren 5,_3, (standard notatlon) one has -ACGTGGGCAATCGAAI. Recall that uracil (U) is not placed into DNA by DNA polymerase.

p1ate, and antiparallel. Reading from the 5, end of the

r;

the

tem_

I strands and second

J

Unwinding of parental round of synthesis (@

ll

5' Lagging strand Okazaki fragments

3'

s'

3,

this deletion is inherited from the father, prader_Wilii sl,ndrome is observed. If the same deletion is inher_ ited from the mother, an enrirely different syrrdrome is observed, termed Angelmann syredrome. The diagnosis can be confirmed by FISH analysis using a p.obe spe_ cific for the l5qll-13 region.

The answer is E: A deletion. The child has prader_ Willi sl,ndrome, which is due to a deletron of a cluster of genes on chromosome 15, on the long arm. When

l2
5'
-r.

3' 4

\s,rGap /n

5'3' - ..:

q3' s',/ .\ f

3'

-: t-i ;--1

i;

'5'

5'

,/

/cap

titting by a repair DNA polymerase

mised. The DNA hehcase is necessary to help ,trbili.. the unwinding of the DNA as the replicarion fork passes through a srrerch of DNA. With reduied heiicase activity, genomic instability occurs, with increased risk of muta_ genic effects and chromosome damage, including chro_ mosome breaks and translocations. These secondary effects lead to the sy,rnptoms observed in the patients. The patients also have a higher than normal risk for various malignancies, due to the increased genomic instability The mutatlon is in rhe BLM gene, which is on chromosome 15. This mutation does not alter, in a direct fashion, DNA poll,rnerase or iigase acrlvity nor RNA polf inerase activitlz Reverse transcriptase is not a normal componenr of eukaryotic cells (it is introduced to cells when they are inlected by a retrovirus).

helicase is defecrive, and DNA replication is compro_

is D: DilA helicase. The chiid has the s)rynptoms of Bloom sprdrome, a disease in which DNA
The answer

l-.
:=

)\A

igure shows one replication fork, moving down the page. As

template must be read in the 3, to 5, direction note how the

13.

T.h: answer is C: Transcription-roupled Dl{A

repair.

::.:--*. rhar must be ligated in later phases of DNA smthesis.

- i.:aki fragments are sprthesized in pleces, moving opposite to ,.: irecrion of replication fork movemenr. it is these Oki)aki frag_

acrively transcribed genes, if RNA polynerase is halted

:lild " exhibiting the sl,rnptoms of Cockayne sy,ndrome (CS), a defect in transcription-coupled DNA repair. Tianscriptlon,coupled DNA repair occurs only on

The

transcribed gene' due to damage to DNA on an acLively transcription this repaircy-rt.* fixes the DNA such that have Cel1s derived from patients with CS can co.rtinue. l lV irradi'ain RNA syrthesi's tn response to
a

is reduced' tion, as transcrlption-coupled DNA repairare at least rvhich did contain thymine cllmers There at bifih' two forms of CS: CS i (or A), the lorm present life' during and CS 2 (or B), one that occurs later in mu[ettons early childhood. The two forms are due to
responsible

reduction

rn genes posis colon cancer, which is due to mutations in DNA mismatch repair' Thls colon *hi.h ur" lnvolr'ed withi'n the tumor does not form large numbers of pollps the other form of lnherited colon canintestrne. as does

from genes thereby reducing the rate of RNA produced

(APC) HNPCC rs also a cer, adenomatous polyposis coh cancer' Defects in base excision repair right-sided colon

ls in two different genes (ERCCS, on chromosome 5' l0' for Ci-A, and ERCC6, on chromosome

A delect in the Wnt signaling an pathwal', which cclnlrols the actron of B-catenin' factor, may play a role in APC' i-porrrr't, lranscription p&cts in reparring double-strand breaks in DNA are linked to br"ast ca,-tcer' Mutations in telomerase would
dJ
r-tot lead to HNPCC

are not is iesponsible for CS-B) The childs symptoms llgase d.,. tt d.f..ts in base excision repair or rn DNA normal tn ln DNA) DNA replication is

Lo an lnabi'1lead to earher ce11 senescence and death due proper length of the chromosomes' ity ro marntain the

(sealing nicks polymerase.

17

fhe answer is B: lncrease in

DNA

mutation

rates'

p53

capability of DNA these children, as i.s the proofreading

and is a protein r'vhich scans the genome for damage' of when damage rs spotted, it i'nd'uces the s1,-rthesis

g.*,
is B:

14- f he answer

fopoisomerase' The quinolone family

oNi

-urr-uliun

to which break the phosphodiester backbone

the gy.ur., li'hicliis the bacterial counterpart ol These are the enzyrnes topoisomerases
al1ow

is unwinding relief of torsional sirain as ihe DNA helix Through an inhibrtion to a11ow replication to proceed is j'nhibited' of gyrase, DNA replicaiion in the bacteria death Since the topoi*t-ti.ll t.ud, to bactetial cell drugs' there is no somerases are not affected by these poll, nerase effect on eukaryotic DNA syrthesrs DNA (the bactena have DNA o O ,r-ttql-,. to eukaryotic ce11s poly-.rur., l, II, oi III) DNA ligase' primase' and drugs The DNA hellcase are not targets of this class of the DNA strands helicase is the enzymt *hlttt a11ows to unwind; however, it needs to work wlth g1'rase (or topoisomerase) such that the tenslon created b1'

s1'nthesis of genes t"he cell cycle. p53 will also lead to the the daminvolved. i.n reparnng the DNA damage Once resume its passage age has been reparred, the ce1l r'u'ill cannot be repaired' thro,-,gl-t the cel1 cycle' If the damage will be imtiated in the celi lf p53 rs missapoptlosis lost' damaged ,r-tg,'o, mutated such that its functions are pire *rtt be replicated, and at times' the replisome will increase make errors repairing the damage This will

*t'rt.t

"otil1

sttp the

ce1l from.

conttnuing through

theoverallmutatlon*ttofthecellsuchthateventually cell promutations lvill appear in genes which regulate a cancer wiil develop p53 ls not involved liferation. and in Wnt signaling or activation of MAP kinases Func-

the lack of tlonal p53 can increase apoptotic events' but decrease the frequency of apoptosts in p:: lvill actually telb. This proteln is also not i'nvolved tn transcriptionr'ouPled DNA rePair

unwinding can be relieved'

:tf,, fhe ansuuer is E: The need for a primer' DNA polyDNA'

t5:

fhe

answer

is G. The drug is

3'-deoxyadenosine

Thls nucleoslde wlll be actlvated when

triphosphate leve1), and

lt

enters

cel1s

will be recognized by t," ,fr. incorporated into growing RNA nNA potl'rr.rur. u"d goup' Since the compound lacks a 3'-hydroxy
chains. phosphodiester RNA syrthesis is terminated, as the next This drug is not recognized cannot be created'

merase requlres a primer in order to s1'nthesize RNA' syntheThe pnmer is provided by a smal1 piece of RNA sltrsired'by DNA primase (an RNA-po11'merase) a smal1 piece of thesis does not requi're a primer. Once to add RNA is s1T-rthestzei, DNApolyrnerase wr11 begin Later' durdeoxyrtbonucleotides to the end of the RNA w11 come ing ONA replication, another DNA polymerase

bond

2'-hydroxy fry ,Ne po11'merase because it contains a will only recogni'ze nuclenro.,r, ur-ti dNA potl'*erase ;ii." which lack 2;-hydroxy groups This drug does it will not not resemble IRNA, or mRNA, or rRNA' so protein slnthesis Protein slnthehave a direct effect on mRNA is present sis may be decreased, however, if no

RNA bases alJng and i.*ou. the RNA, replacing the syrwith'deoxy-ribonucleotldes However' as rnitlally While uracil thesized, Okazaki fragments wiil contarn of rytosine can produce uracil' this is the deaminatlon than RNA much more frequent in the more stable DNA (tt cannot be easily converted to uracil in DNA

inhibit to translate. However, the drug cloes not directly the mechanisms of translation'

Th).'mine not contribute would require a demethylation), and does h Okazaki fragments Mismatch repair to uracil content

16

repair' The The answer is A: A defect in Dl{A mismatch nonpolypnit"r-t, o concerned about HNPCC, hereditary

(whlch form does not operate on DNA:RNA hybrids pol1'merase when the primer is syethesized)' and DNA uracil, so it would not make the type does not recognize DNA' of mistake ln which uracil were placed into

\ \
DNA Structure, Replication, and Repair
The answer is

27

(: (reation of a fusion protein not normally

found in cells. The patient exhibits the Phiiadelphia chromosome (a translocation between chromosomes 9 aad22), which creates the bcr-abl protein, a fusion pro;ein of two normal ce11ular proteins. The ab1 protein is a :rrosine kinase; when the gene is moved from chromosome 9 to 22 and putunder the control of the bcr gene, :i rs mise4pressed -zifid constltutlvely actlve and leads to :ellular growth in the blood ce1ls in which rhe protein = expressed. The patient has chronic myelogenous leu<emia (CML). Since this is a domlnant activity there = no loss of a tumor suppressor gene. The transloca::..n does not interfere wlth DNA repair, so there is no :irect increase in mutation rate due to this transloca-rrn. There is no loss of tyrosine kinase activity (there is

actually a gain of activity), nor is there gain of a serlthr kinase (as ab1 is a tyrosine kinase).

!#i ne answer is B: Ribonurleoside triphosphates. DNA

,+?.t:.t::,:

s),nthesis requires the syethesis of prlmers upon which

deoxyribonucleotides

will be added. The primers

are

composed of ribonucleotldes (DNA primase is a DNA-

dependant RNA poll,rnerase), and the sclentist forgot

to add NTPs to the reactlon mixture (deoryribonucleotides are not recognized by DNA primase, and cannot be used to slrrthesize a primer) Reverse transcriptase is not required for DNA s),nthesis from a DNA template (as in this situation). Sigma factor is a required factor for bacterial RNA slnthesis; it is not a required factor for eukaryotic DNA sy,nthesis.

r
5

n
E

L
]E

I.
m
@

Chapter 4

RI-{A Synthesis
Thr cytestio*s in this chapter Gr{: a{g{t1-,izttl {r.l}*z t tkt prorcss oJ rrnrisrr i;-:fian and disrrisrs *:zd yroblmzs associstcd u,illt tr**sct iytiott. Rrgl.l;:'tia* af gme exlst"cssiottwillbe {o"t,tlTdinnrct-e detsil in
Chapter 6.
.o\
-t'
-o'@

-(o"
^ao' Y

s,v

^9

"e

QUESTTONS
Select the single best answer.

An IV drug user is resred, and found positlve, for infection by HIV If the patient is only placed on one antiviral medication, liral loads wlli inirialiy be reduced, but wr11 then rapidly increase. The resistance to the drug occurs
due to whrch of the following? (A) Lack of error checking in DNA polymerase (B) Lack of error checking 1n RNA pol;,rnerase

(C) Lack
celis

of DNA repair enzyme sysrems in HlV-infected

(D) Incorporation of uracil in the RNA genome of HIV (E) Incorporation of th)..rnine in the RNA genome of HlV A researcher needs to prepare RNA for Northern blot analysis. Initial experiments using total RNA produce
no signal when the experiment is completed. A method to increase the sensitivity of the assay would be to do rvhich of the following ro the total RNA sample? (AJ Separate by srze on agarose ge1 electrophoresis (B) Run the RNA through an oligo-dT affinity column (C) Run the RNA through an oligo-dA affinity column (D) Separate by stze on polyacrylamide ge1 electrophoresis

(A) Codon degeneracy within the genetic code (B) Tissue-speclfic postrranslatronal modificatlons @) Tlssue-specific alternative sphcing of the primary
transcript

(D) Polyadenylation is lacking in cerrain

tissues

rissues

(E) Differences in locarion of 5'-cap formation in the

An lndividual having B-thalassemia minor exhibits rwo bands on a Northern blot using a probe agalnst exon I ol B-globin. The smaller band is of normal size and "heavler" than the other larger band, which consists of
approxlmately 247 addittonal nucleotides. One explanation for this finding is which ol the following? (A) The presence of a nonsense mutation in the DNA (@) A mutation which creates an alternatlve splice site (C) A lack of capping of the mRNA (D) An extended poly-A tarl (E) A loss of AUG codons

(E) Perform a phenol exrracrion of the total RNA

A researcher has obtained an antibody to cytosolic protein X and runs a Western blot uslng as samples a variety of tissue t1pes. The resuhs of the Western blot are shown below. A potential interpretation of the results is which of the following?

careful analysis of ce1lu1ar components drscovers short-lived RNA species in which an adenine nucleotide is found with three phosphodlester bonds (linked to

28

RNA

Slnihesis 29

the 2' , 3', and 5' carbons). This transient struciure is formed during which of the following processes? (A) mRNA cap formation (B) mRNA polyadenylation (e) Splicing of hnRNA (D) Transcription of m.itroRNAs (E) Transcription ofiRNA A patient suffering from chills, vomiting, and cramprng rvas rushed to the emergency department. He had eaten r,nld mushrooms for dinner that he had picked earher in the day. His sl.rnptoms are due to an inhibition of u,hich
of the following enzymes?

(A) RNA polymerase I (B) nNA polymerase II (C) RNA polymerase Il1 (D) Telomerase (E) DNA primase
$estions 7 and 8 refer to the figure of active transcription
:

"1o*'. ,, RNA polymerase

(A) (B) (C) (D) (E)

Ribonuclear proteir-r complere

D\A

polunerases

Carboh,vdrates

tRNe complexes Peroxisomal proteins

Hybrid
RNA-DNA
Direction of transcription
1S

Ribonucleosidetriphosphates

Addition of
ribonucleotides to growing end of transcript

t0 When first dlscovered, introns were not thought to code lor a functional product. Recentiy, hower.er, introns
have been found to code for products that can regulate the expression of a large number ol genes. This regulatron occurs at which stage of gene expression?

,rimary
,
,.,

he message identical strand is best represented by

hich letter on the diagram?

in the
re 3' end of the newly s1T-rthesized RNA is best repre..nted by which letter on the above diagram?

(A) Transcriptlon of mRNA (B) Posttranscriptional processing (C) Export of mRNA into the cytoplasm (D) Ribo,ome biogenesis

(f)
t'l

oegraciation of the mRNA

bits two
1St exon

size and of 'nsists

.-. 22-year-old woman (see the frgure below) sees her ::r sician for a variety of complaints over the past year.

re exDlaEl re

:ese include fevers that come and go, fatigue, .loint and stiffness, a butterfly rash on the face, sores .,-. 1er mouth, easy bruising, and increased feelings of
-..rn

A researcher, while studpng a liver ce11 line, found the foliowing anomalous result. He u.as studying protein X production withrn the liver ce11. \i,'estern blot analys.is
using a polyclonal antibody shor.ved a normal size, and amount, for protein X. Enzyme assal's demonstrated normal levels of actlvlty for protein X. Northern blot analysls, however, pelded two bands of equal intensity: one the expected size and the other 237 nucieotides

DNA

,.irery and depresslon. A diagnostic blood test is hkely

i:1ow autoimmune antibodies drrected against which -..; ol molecules?

rlice site

discover.

r,ucleotid;

,linked

tc

3A

Chapter 4

longer. One possible explanation for this finding is

which of the following? (A) A nonsense mutation in the DNA (B) A loss of an intron/exonjunction (C) Inefficient transcription initiation (@ Loss of a transcription termination site (E) Gain of an alternative splice slte

in an lntron of the B-globin gene. How does such a

mutation lead to this clinical finding? (A) A microRNA is produced, which is rargered againsr

the B-globin mRNA, thereby reducing B-globin production (B) Creation of an akernative splice sire, such that
B-globin levels are decreased

.l.L

(C) Creation of a new rranscription initiaLion site, such

A young child of Mediterranean parents was brought to the pediatrician due to lethargy, tiredness, and pallor. Blood analysis revealed a mlcrocytic anemia, ,although iron 1eve1s were normal (see the figure beiow). What test should be run to determine that the chiid has a variant of thalassemia? that the mRNA for B-globin is now our of frame

(D) Creation of a stop codon in the B-globin mRNA (E) Ellmination of the polyadenylatlon signal, thereby
reducing B-globin productlon

,*5.

Consider two individuals, each with some form of thalassemia. Patient X has a deletion of the o, genes on one chromosome but normal expression of all other cr and B genes. This person has a mild form of the disease. Patient Y has a normal complement of cx genes but has a homozygous mutation in the B genes in which an abnormal spllce site is used 80o/o of the time, producing a rranscript with a premature stop codon. Patlent Y has a more severe disease than patient X. Why is patient Ys disease more severe than patient X's?

@ W.tt.*

blotting of the peptide chains in hemoglobin

(B) PCR of the gene for RNA poll.rnerase (C) Western blot of snurps in the chl1d
(D) Western blot of TFIID

(E)
,13

PCR of the gene for y-globin in the child

(A) The ratio of crlB in patient Y it is 1:5 fne ratio of crlB in @ patient Y it is 5: 1 (C) The ratio of ctlB in patient Y it is 1.2:1 (D) The ratio of ctlB in patient Y it is 1:1.2 (E) The ratio of c/B in patient Y it is 1.2:l

parienr X is parienr X is

l:2,

whereas in

l:2, whereas ln

patient X is 2:1, whereas in patient X is 2:1, whereas ln

patienr

is 1:2, whereas

in

An intestinai cell line was being studied for its ability

to produce lipld-containing partrcles. Surprisingly, a mutated vadant of this cell line was unable to do so. Western blot analysis yielded a protein with the same size as apolipoprotein B100. A potential mutation in this cell line, which would lead to this result, is which of the following? (A) Splicing defect (B) Cap formation altered @ nNa editing defect (D) Inefficient poly-A tail addition (E) Promoteralteration
A patlent displays tiredness and lethargy, and blood work demonstrates an anemia. Western blot analysis indicates

.:ie,. Dideo$mucleotides are effective agents against DNA q,mthesis, but appear to have litt1e, or no, effect on RNA s1-nthesls. Thls is mosr likely due ro which of rhe

following?

'""ff"""
HH
A dideoxynucleoside

(A) Lack of a 3'-OH group fB) Lack ola 2'-OH group

IC)

Presence o[ a 5'-phosphodiester bond

significantly greater levels of u-globin than B-globin.

Molecular analysis indicates a slngle nucleotide change

(D) Presence of an N-glycosidic bond at carbon I (E) Factor TFIID does not recognize deoxyribonucleotides

RNA Sl,nthesis

3l

ffi

rffflA i, u J*rrury
(A) mRNA (B) rRNA

rranscriprion factor for the synthe-

t:.i?i:;

sis of which class of molecules?

A man with a bacterial infecrion was prescribed rifampin to resolve the infection. Rifampin does not affect eukaryotic cells due to which of the following?

(A) Differences in ribosome structure between eukary-

tRNa

(D) hnRNA

(E) microRNAs

CE)

ff; ^ 2-year-old, has been diagnosed with a rhabdomyosarcoma and is piaced on chemotherapy, includlng the
drug dactinomycin. Dactinomycin exerts its effects by which ol rhe following mechanrsms?

otes and prokaryotes Structural differences in RNA polymerase between eukaryotes and prokaryotes (C) Differences in transcription factors between eukaryotes and prokaryotes

(D) Inability of the drug to bind ro DNA conrainlng

nucleosomes

(E) Differences in snurp structure between eukaryotes

and prokaryotes -::*,.:, A cell line was derived, which was temperature sensitive for splicing hnRNA. At the nonpermissive temperature, splicing was unable to occur. A potential activity that is murared in the splicesome is which of the following? (A) Ability to carry out RNA sl,nthesis

nindirrg of the drug to DNA, thereby blocking RNA syrthesis (B) Binding of the drug ro ribosomes, thereby blocking translation (C) Binding of the drug to transcription facrors, thereby blocking RNA sprthesls (D) Binding of the drug to RNA poll,rnerase II, thereby blocking RNA sl,nthesis (E) Binding of the drug to DNA, thereby blocking DNA
sy,nthesis

(B) Ability to carry out DNA s),nrhesis (C) Loss of 3'-5' exonuclease actlyity

Loss ol endonuclease acl ir,iry Loss of ability for transcription-coupled DNA repair

(E)

ANSWERS

'{ fhe answer is B: Lack of error checking

in Rl{A poly'

merase. As part of the life cycle of the virus (see the figure below), the RNA genome of the vi'rus is conuerted to DNA, which integrales randomiy into the

cannot check its work and cannot fix errors when a mismatch is made. The accumuiated effect of these errors increases the mutation rate of the virus much more than organisms containing DNA genomes' Since the enzyme that creates the viral DNA is reverse [ranscriptase, which also has no error-checking capability'

host chromosome. Host cell RNA polymerase lI transcrlbes the viral DNA, producing viral RNA' which is translated to produce viral proteins' and which ls RNA aiso utilized as the genome for new virai particles activpo11'merase does not contain 3'-5' exonuclease DNA polpnerase does), so RNA poil'rnerase ity i*hich
then
Retrovirus Particle

the risk for mutations is greatly enhanced' DNA polymerase does check its work but ls not used in the viral life cyc1e. The DNA repair enz:ptes are not altered by HIV infection. Uracil is a normal component of the viral RNA genome, whereas th)''rnine is no[' but neither of
these facts results

in an increase in mutation rate'

each containing ,r,'l reverse transcriPtase'

An oven lew of the retroviral life cycle'

affin' The answer is B: Run the Bt{A through an oligo-dT

conditlons (through a reduction of salt concentration)

can then elute the mRNA specifically from the column'

ity tolumn. Mature mRNA from eukaryotes has a poty-A u11, which is added posttranscriptionally by

(see the figure below for an overview ,"rr-O polyrnerase mRNA s)-nthesls) The poly-A tail will of ."Utyoti.

hybridize to the ollgo-dT on a column, thereby allowing the mRNA to bind to the column and other types of"BNa to pass through the coiumn' Altering the salt

Phenol extraclion is required for nucleic acid isolation' but it i.s not specific for mRNA. Electrophoresls, either by agarose or polyacrylamide, will separate nuclelc acids isolation' An sLe but does not by itself lead to mRNA oligo-dA column will not hybridlze to the poly-A tail of
mRNA.

Nucleus

-cap

5'-Cap

i5
s'./

-r'
DNA

I t

: t\

,S'-CaP

poly-A
hnRNA

Nuclear envelope

)-----* 5 I/\ poly-A Nuclear "'/ pore

mRNA

RNA SirLthesrs
The answer is C: Tissue-specific alternative splicing of the primary transcript. Tissue-specific akemative splicing of

33

- -,e primary transcript can give rise to a number of dis_ ,-:r.r mature mRNAs, each of which gives rise to a vanant .he parent prorein tbur all separale proreins in their own -lhr) (see the figure below). posttranslarional modifica_ .-rns are made primanly to membrane bound or targeted

sequence; it

speclfy the same amino acid in the proteins pnmar)wll not alter the sequence o1 amino acicls in

proteins, but not cytosoiic proteins. polyadenvlation and cap formation do not alter the reading frame.of the protein and are required to fully mature the mRNA. Codon degeneracy wrll a11ow a number of different codons ro

the fina1 prorein as will alternative splicing.

Cleavage site for short transcript 5'-splice site 3'-splice site

Cleavage site for long transcript

lnitial membrane-bound

Stopcodonl I Transcription antibody I I

I

Stop codon 2 Secreted antibodv

V -:rg RNA transcript

Stop codon 5'-splice site

-.........-r

t
Short RNA transcript
Stop codon

Stop codon 2

3'-splice site Poly-A

L________l_-__l

lntron removed
by splicing

-RNA
Stop codon 2 Poly-A
I

rranstatton

+ ',lembrane-bound antibody (IgM)

I Transtation

+
Secreted antibody (IgD) COOH
_t________J

L_

Terminal hydrophobic peptide

Tu,o drfferent anribodies

Terminat hydrophilic peptide

as a

COOH

(lgM and IgD) procluced from the same initial RNA transcnpt

result of alternatir.e splicing.

::ted into the mRNA product, producing a longer than :ormal mRNA This is an infrequent event, however,

The answer is B: A mutation which creates an alternative splice site. The patient has cleveloped a mutation r an rntron whlch acts, oniy a small percentage of the .-me, as a splice donor site instead of the normal site at -ne intron/exon boundary Thus, when this sire is utr_ -, -ed by the splicesome, a piece of the intron is incorpo_

If the polyadenylation

srgnal were murared, then the

overall mRNA size would be larger, but there would not be two different proteins produced. Slnce the parrenr has a B-thalassemia, defective B-globin protein ii being produced lrom the larger mRNA. Loss of methrontne codons mll affect translation, but not transcriptron.

rai band. A nonsense mutation in the DNA will not ,:ifect rranscription (although ir does affect rhe proreln :roduct made from the mRNA) The lack of a cap would :esu1t ln an unstable mRNA that perhaps woulcl not be -ranslated but r,vould not significantly change the size of :ne mRNA. Poly-A polymerase adds the poiy-A tail and ,i'ou1d add the sarrre size rail to borh specres of mRNA.

sred mRNA band on the gel is darker than this

,:s.judged by the finding rhar rhe density of the normal

abnor_

The answer is C: Splicing of hnRl{A. As seen on page 34, an adenine nucleotrde in the micldle of rhe intron rs a required component for sphcing to occur, and the sugar residue attached to this adenine is rnr.oh.ed in three phosphodiester linkages; the normal 3, and 5, ancl also 2' to the sphce site. The resulting structure resembles a lariar :uch a struclure does nor lorm dunng capping polyadenylation, or the normal transcription of genes. It is unique to the splicing mechanism.

34

Chapter 4

lntron--_------1 , .;
tJ

the template for RNA synthesis (which is designated as C in the flgure).

as

ffif,t
First

a*Effi
U2,
Second cleavage
site

cleavage
site

Ul1

The newly rynthesized RNA strand ls made ln the 5'-3'directi.on, so letter A represents the 5' end of the newly sytthesized RNA and letter D represents the 3'end. The DNA template is being read in the 3'-5' directlon (letter C would represent the 3' end of
The answer is

D.

the template strand).

s*FffiE

The answer is A: Ribonutlear protein complexes. The woman has lupus, an autoimmune disorder. One class of antibodies developed is agarnst the snurps, smal1

ribonuclear protein complexes, which are involved in mRNA splicing. Autoantibodies are not developed against DNA polymerase (although antibodres against

ffi
Lariat
u-2

DNA are often found), carbohydrates, IRNA complexes, or peroxisomal proteins.

to

The answer is E: Degradation of the

mRllA. lntrons

have

Lariar formation during spii.cing, showing the required tntronic adenine nucleotrde wrth three phosphodiester bonds.

been shorvn to contaln genes for microRNAs, which are processed to smal1, interfering RNAs, which can regulare gene expression either by binding to and rnitiating

The answel is B: RllA polymerase Il. The patient has ingested G-amanitin, a toxin that, at very low concentra-

tions, inhibits RNA pol1'rnerase II and blocks the transcription ol single-copy genes. RNA polltnerase I and III are more resistant to the effects of amanitin, and this roxin has no effect on telomerase or any type of DNA pol1.merase. The rnability to s1'nthesize new proteins in all cells leads to the symptoms observed. The structure of amanitin is shown beiow. Amanitin porsonlng lnltially causes gastrointestinal disturbances, then electrolyte imbalance and fever, followed by liver and kidney dysfunction. Death can fo11ow 2 to 3 days after ingestion'
OH

degradation of a particular mRNA or by binding to a particular mRNA and blocking translatlon of the mRNA' These sma11 RNA molecules do not affect the transcription of the target mRNA, nor posttranscriptional processlng (capplng and polyadenylation). They also do not affect the export of mRNA into the cytoplasm, nor do they alter ribosome bi.ogenesis. As an example, the mlRl7-92 cluster encodes seven microRNAs and resides within an inlron of the C13 or F25 gene on chromosome 13. These miRNAs are upregulated in lung cancer, and. may contnbute to the progression of the disease by downregulating their target genes.

H.C CH_CH,OH " \,/ tl^l oc

HN

CHH tl

CH-CO-NH

-c H,clr

co

NH- cH2-co

Pre-miRNA complex

f ?* 'otTcHoc
I

Tn."r, o=1^P/'"",\','-?^$r,cr.
NH

-f)

-CO -CH-NH-CO -CH2o-Amanitin

?"

?o
NH

Pre-miRNA complex

H2C

CONH2

An overvlew of microRNA lranscriplion, processing, and mode of

action. The mrRNA genes are lranscribed ln the nucleus, processed to a pre-mi.RNA, and exported to the cytoplasm. ln the cytoplasm, the pre-miRNA ls further processed by an RNase (Dicer), and the resuliant double-stranded mIRNA forms part of the RISC (RNA-induced silencrng complex). A strand selection separation process occurs rhat allows recognition of the appropriate mRNA to ablate (either by nuclease degradation or lnhtbition of translation)

'

The answer is

is the one that has the same sequence of

The message-identical strand of DNA bases as the mRNA product (with the exception that when there is a U in RNA, there is a T in DNA). Thus, this is the strand that is complementary to the strand that is berng used

B.

RNA S1'r-rthesrs 35

il

u
:L

t\'e
-..re

The answer is D: Loss of a transcription termination site. One of the genes that encoded protern X had a ::- -.--rlion at the transcrlption termination slte, which .:-...:1ed the mRNA to be transcribed into a longer form -l- nucleotides longer). The reading frame of the ..-1l{-\ u'as intact, as was the slart and stop codons, so , -: :rolern produced lrom this lengthened mRNA rvas :- --:nal. Il a nonsense mutation had been created in gene :. :runcated, ilkeil'rnactive, protein w'ould hal'e been -:-.iuced. The loss of an intron/exon junction would ,, .:: the sphcing paliern of the mRNA, and r'vou1d most ..-:-r- aiter the reading frame of the protein and create ' ::,-lfunciional protein. Another possrbility is that the :i Lri an intron/exon junction would produce an elon:,,::c1 protein (due to intronic DNA being transcrihed as r,:-- rri the mRNA), with a concomitanl loss of aLtl\-il\ potentialll'lead -r-..-lir-rg an aiternative sphce site r'l'ould -'io forms of the flna1 protein being produced, r'et ---'. one is seen by Western blot. inelficlenl transLrlp:- rnittarion would not produce tr,vo distinct mRNAs
The answer is A: Western blotting of the peptide chains an in hemoglobin. Thalassemias are the result

opposite would be true lor a B-thalassemia). As manr cr-thalassemias are caused by gene deletions. FISH might be another way to determine this condition.

using a probe specific for the u-globin gene lr'lost for B-thalassemras are not due to gene deletions PCR y-globrn (feta1 Hb) or RNA polymerase will not address
an imbalance
Simi1ar11',

Western biots of snurps or TFIID lvill not address an imbalance in.synthesis (if there r'vas a problem r,vith snurps, all RNA splicing rvould be affected, not just the cx or B-globin gene; similarl)', il TFIID w'ere altered, all mRNA synthesis would be altered). Clinrcal labs will also use hemoglobin electrophoresls to quantitate the levels of globin chains in a patient. The illustration used in the question \,vas obtained from a piltienr
rr

in cr- and/or B-globin chain

synthesis.

ith B-thalasscmra.

13

The answer is

12

lng
-.r a

ol

:':-li.Lnce

in the synthesis of cx- and B-globrn

genes.

\,\.

,in, 'r

proI no[

-::rs rvere to occur, a Western blot anal)'sis of the cr : :. B chains u,-ou1d show a difference in the amount . . .,'.ih in the red blood cells, suggesting erther an

,r do
n'riRi10e5

-.r 9-rhalassemia (in an cr-thalassemia, one rvould ::: -.ss cx chains or variants of cx, chains being pro. ,.-.-d. as compared to just one, normal B chain. The

,-

editing defect. The intesrine uontains an R\A ecliting complex thai alters one base in lhe apo B10tl n-rRNA. ri-hich creates a stop codon, such that u-hen the r.nR\A is translated. protein sl.rlthesls stops alter 48?o ol rhe codons have been translated This is a unique tlpe of posttranscriptional n.ioclifrcarion. The imtial rranscnpts lor both apo B-18 and apo 8100 are the same. Mutations thal alter splicrng. cap fr)rrn3tlon, or polyadenylation would not prodr-rce the full siue protein in place of apo B48. Mutations ln the promoter u'ou1d alter initiation of transcription, but not rhe end product fonrred. RNA editing in this case is shourL 1n the figure
RNA

(:

belolv

omolNCCI,

B-apoprotein gene
C

:se

by Liver

1NA )avage

Transcription
and

lntestine

I
t5u
A
I

mRNA 5'

g' ^..-r\-n-lCr.-n -4=n

RNA editing

5'a'A--r1-Lu-t n--n--n
(Stop codon)

Lranslation

{i (
ricer

Apo B-100
4,536 amino acids

*f.
amino acids

ApoB-48

-2,152--]'

nplex

mode ol
rcessed to

of -l : l3: li-r intestinal cells, RNA editing conr.erts a cytosine (C) to a uracil (U), producing a stop codon. Consequently, the B-apoprotein Apo B,lB is 43% rhe size of apo Bl00, ihe product syrthesized in the liver from the -: ls lapo B4g) contains on\,Z,irZ amino acrcls.

:i:ism, the
',he resul-

,',:d rvhlch was not edited at the RNA 1evel.

r-rnduced
.SS
OCCUI:

r rei[her b\

Chapter 4

14

The answer is B: Creation

such that p-globin levels are decreased.

of an alternative splice site, if an intronic

mutation creared an alternative splice site, the splice_

some wouid utilize this site for splicing a cerrain percenLage of the time, forming an mRNA that would nor code

poll,rnerases are labeled TFIIx or TFIx, respectlvely MicroRNAs are s)Trrhesized by poiymerase IL hnRNA is the precursor for mature mRNA, also syrrthesized,by RNA pol;.merase ll

for funcrionai B-globln protein. This would lead to

t8

reduction in B-globin s),nihesis relarive ro u-globin syrr_ thesis, thereby crearing a B-thalassemia. If a microRNA were creared which targeted the B-globin mRNA, then there would be a drastlc reduction in B-globin sl,rlrhesis as all B-globin mRNA would be targered for desrrucrion, which is not observed. Since the introns would be nor_ mally spliced from the marure mRNA, the creation of a transcription initiation site would have no effect on the mature mRNA. .Simi1arly, the creation of a stop codon in an intron wouid have no effect on the mature mRNA. The polyadenylation signal is not found in introns, so rhe mutation could not be at this locatlon within the gene.

of the drug to Dl{A, thereby blocking Rl{A synthesis. Dacrinomycin binds to DNA and blocks the ability of RNA poll,merase ro transcribe genes, thereby blocking rranscriprion. The drug does not bind specifically to ribosomes, transcription factors, or RNA poly.merase II. It also does not interfere with DNA sprthesis.
The answer is B: Structural differences in Rl{A polymerase

The answer is A: Binding

t9

between eukaryotes and prokaryotes. Rifampin binds to prokaryotlc RNA polymerase bur cannor bind (due to the different structures of the RNA poll.merase between

15

The answer is B: The ratio of o/B in patient X is l:2, whereas in patient Y it is 5:1. An individual with the genotype of patient X is producing 50% normal cx chain (two normal genes, rwo deleted) and 100o/o B-giobin for a ratio of 1:2 of o, to B(each chromosome 11 contains

prokaryotic and eukaryotic cells) to eukaryotic RNA drug does not bind to ribosomes. DNA, snurps, or transcription factors. Snurps are not present in prokaryotic cel1s.
pol1.merase. The 20

two G,-genes, for a total of four o,-genes per ce11). Such a ratio would lead to little, if any, clinical symptoms. Patient Y is produclng 100% cx chain and 2Ook chaln B for a 5:t rario of cr to B. This ratro is big enough to lead to clinical s)rynproms. Recall, the major biochemical problem in thalassemia is the imbalance in synthesis of o, and B chains, leadlng to nonfunctional s, and Fr tetrame.s forming from the excess chains produced. The other ratios presented as answers do not represent the mutations present in the patients.

The answer is D: Loss of endonuclease activity. The act of splicing requires the breakage of internal phos_ phodiester bonds, which is the job of an endonuclease. Spllcing does nor require new RNA sy,nthesis, DNA s1mthesis, error-checking (the 3'-5' exonuclease actlvity), or DNA repair. The process of spllcing is shown again
below.

ffi':=. /
First

a4-a*ffi
Second cleavage
site

l6

group. RNApolymerase is looklng for substrates that contain a 2'-hydroxyl

Theanswer is B: Lackof a 2'-0H

cleavage
site

group (reca11, DNA poil,merase urilized dNTps, whlch normally iack a hydroxyl group at the 2' position). As this substrate lacks a 2'-hydroxyl group, therefore the blnding affiniry of this drug for RNA poll.rnerase is very Iow, such that the likelihood that this chain rerminator will be incorporated into a growing RNA chain is minimal. DNA polyrerase, however, utilizes substrates lacking a 2'-hydroxyl group, and can bind and utilize
this substrate.

17

tRl{A. RNA polymerase III requires transcription factors to bind to promoters, which are
The answer is C: labeled as TFIIIx, where x is a variable letter. pol)..rnerase wiil sl,nthesrze IRNA molecuies and 55 rRNA RNA poll.rnerase II sl,nthesizes mRNA while RNA polymerase
111

+A-..-+-Aii
.. u2

,."

u+lo

'.

u5

ffi

syrLthesizes primarily rRNA. Accessory factors for the

Larlat formation during splicing, showing the required intronic ade_ nine nucleotide with three phosphodiesrer bonds.

Chapter 5

ffr#E*Eee %yeaFe*rus
:,tt,S{r::'
',..ii c
tz';

{oit{s th* b*sics

*.{

prrttein 5';stl:-i:sis

i. cr:rj E1*1i, d:"!fuy{n',ccs ls*wtczt ysr*l*.r",'':iir '.,.]:a*'*tir ira*s1*tit>tt *r* *xyl*it,:d ia', ti-.i
t:ittii-.i-*tir,s. A ttttmzbry *f srutstirstls {rJii !1'{ :f . tG tiir gtwr.tit: :;itl*, v,l'zi#z ls ye-*t;;t':i,
?-

intluriizzg

str r sz; si*

ti*n*l

wi: Gzf,-

synthesrs. Analysis of the mRNA produced at the nonpermi.ssive temperature indlcated that a key structural

feature, normally present on mRNA, was missing Such a structure is most likeiy r,vhich one of the fo1lowlng?

iL

J":.

::
;

e 5-1.
Base

(A) (B) (C) (D) (E)

Intron-exon secondary slructure

Pseudoundine The 5'cap Thymine The poly-A tail

-t

Second Base

Third Base

UCA
Phe
Phe

G
Tyr Tyr

(3',)

Ser Ser

cvs cys
Stop
Trp

Leu Ser
Pro

Stop
Stop

Arg His Gln Gln

Under conditions ol active exercise, proteln sp-rthesis is reduced in the muscle. Under these conditions, which aspect o[ translation is jnhiblted? (A) InabrllLy to initiate translation (B) lnabllity to elongate during translation (C) Inability to terminate translation (D) lnabllity to sy'nthesize mRNA (E) lnabllity to produce rRNA

n
Leu Leu lle lle

Pro

Arg Arg Arg

Ser

A young child exhibits the lollorving symp[oms: Coarse facial features, congenital hip dislocrtion,
inguinal hernias, and severe developmental delal'. These symptoms are lully evident at the childs age of I. Ce11u1ar analysis demonstrated the presence o[ inclusron bodies within the cytoplasm of liver cells. The inclusion bodies are the result of which of the
following?

Asn Thr
Lys Lys

Ser

Arg Arg
Glv Glv

Met

Thr

Asp Asp
Glu Val

(A) Enhanced lysosomal enzyme activity

btv
(]tv

Ala

Glu

tB) Reduted lysosomal enzyme activity (C) Enhanced peroxisomal enzl,ne activity (D) Reduced peroxisomal enzyme activity tE) Enhanced protein secret.ion

tL ESTHONS
:{:i!t 1
ttre single best answer. --- researcher has discovered

A eukaryotic

cel1 1i.ne contains an aberrant, temper-

a temperature-sensitive -:-1 ltne that displays an overall reduction in protein

ature-sensltive ribonuclease that specifically cleaves the large rRNA molecule into many pieces, destroying its secondary structure and lts ability to bind to ribosomal proteins. This cel1 ltne, at the nonpermissive temperature, has greatly reduced the rates o[

37

38

Chapter

protern synthesis. This rate-limiting step is which of

the follor'ving?

(A) Activation of protein kinase A (B) Activation of an elongation factor for translatlon

(A) Initiation

rCi Gl)cosylation ola C protein

(D) lnhibrtion of protein kinase A (E) lnhibition of an elongation factor for translarion

(B) Termination (C) Elongation (D) Peptrde bond formation (E) IRNA activation and chargrng

A 2-year-old girl exhibits a very high fever of sudden

onset and compiarns of a stiff neck. Physical exam reveals

5 A cell contains a mutated

alanine-tRNAnl" s1,-Lthelase that recognizes glycine instead of alanine as ils subslrate. The anticodon of the IRNA recognized by this enz)-rne rs IGC. When the ce11 translates the fol1ou-ing portion

positive Brudzinski and Kernig sign and petechlae on the extremities. The pediatrician, in additron to rushing the child to the hospital, prescrlbes a drug that blocks
a

of an mRNA molecule (presented in frame beginning with the 5'nucleotide), u'hai mll be the amrno acid sequence of the proiein produced lrom this stretch of
mRNA?

prokaryotic peptide bond formation, even though it can har.e serlous side effects. That drug is which ol the
following? (A) Rifampin

-AUGGCGGACUCGGCUAUG-

(A) M-G-S-D-G-\,1 (B) M-A-D-S-G-\'l (C) N4-A-D-S-A-M (D) NI-A-S-D-A-M (E) \1-G-D-S-A-M
Questions 6 an'dT refer to the following case. A 3-year-old boy, whose parents did not immunize him due to fears of postimmunization side effects, exhibited fever, chills, severe sore throat, lethargy, trouble breathing, and a husky voice. Physical exam indicated greatly enlarged lymph nodes, an increased

(B) Rapamycin (C) Chloramphenicol (D) Cycloheximide (E) Puromycin

A Russian youngster w'as prescribed erythromycin for a bacterial infection, but he developed hearing loss due to use of this drug. This occurred due to r'vhich of the following? (A) lnhibltion of mitochondrial protein s)nthesis

heart rate, and swelling of the palate. A picture of the boy's throat is shown below

(B) Inhibrtion of mitochondrial RNA s)-nthesls (C) Inhibition of mitochondrial DNA replication (D) Weakening and tearlng of the eardrum (E) Increased neuronal signaling in the inner ear

While investlgating struclure-function studies

in

membrane transport protein, a researcher discovered a single nucleotide mutation that 1ed to the loss of a key u-helical segment of the protein. The mutation that 1ed to this fi.nding is most like1y which of the following?

(A) CUC to CCC (B) GUU to GCU (C) UUG to CUG (D) AUC to GUC (E) GUG to UUG

tt
The throat is du11 red, and a gray exudate (pseudomembrane) is
present on lhe uvu1a, pharF-rx, and tongue.

An adult male is diagnosed with a typical pneumonra

His physician prescribes clarithromycln, whrch is specific for prokaryotic cel1s. Which of the following best
explains the mechanism of prokaryotic specificity?

A necessary cofactor for allowing these symptoms to appear in the chlld is which of the following? (A) ATP (B) NAD. (C) FAD
(D) Acetyl-CoA

(A) The drug brnds to the 50S ribosomai subunit of

bacteria and lnhibits f-met-tRNAr bindrng

(B) The drug binds to the 30S ribosomal subunit of

bacteria and blocks initlation o[ protein s)'nthesis

(C) The drug binds to the 50S ribosomal subunit of

bacteria and blocks translocation

(E) UDP-glucose

(D) The drug binds to the 30S rlbosomal subunrt of

bacteria and blocks peptlde bond formation

The molecular mechanism responsible for these physica1 obsen ations in the boy rs which of the following?

(E) The dr-ug binds to both ribosomal subunits and

prevents bacterial ribosome assembly

Protein Sy,r-rthesis
-a-

39

:etient underwent a kidney transplant and among

-r-. many drugs she received posttransplant was ::::mvcin. Rapamycin aids ln preventing an immune
r:-.3elrrse to the transplant via which of the following -::hanisms?

(A) 1 (B) 2

(c) l
(D) 4

(E)
,+7

--: I :

The drug inhlbits ribosome subunit assembly The drug inhibits cap formarion The dmg specifically inhibits RNA poll,rnerase The drug lnhiblts rnitiation of protein s)'nrhesis

III

A young boy has edema, a protruding abdomen, and very thin arms and legs. The edema has at lts origins
which of the following?

The drug inhrbits antibody-specific transcrlption

factors from binding to DNA

-i.

-. see a very sick patient (vomiting and bloody -'.rrhea, dehydration, and mental status changes) in
emergency department, who, you are told, was an

(A) Lack of muscle protein slnihesis (B) Lack of liver protein s),nthesls (C) Lack of intestinal protein s)-nthesis (D) Excessive water productlon due
hydrolysis (E) Excessive water production due hydrolysis
l:-:a::i::: '

ro

protein

::rateur chef trying out a new creation in which

to

triglyceride

he '.;anted to experiment wrth the extracts of castor beans.

ihs
i-ie

persons symptoms are all due ro which of RNA polymerase I

of RNA poll.rnerase

of

the

i..11orving?

,j.*.

-\) Inhibition B) Inhibition

A patient taking lovastatln and Zetia@ lor elevated cholesterol was found to produce lower levels of glycosylated proteins. This is most 1ike1y due to the unintended consequence of blocking the sgrthesis of which of the following compounds?

lI

he

.C) Ribosomal inactivation by covalent modificatron

D) Ribosomal disassembly due to covalent modification .E) Inhibition of amino-acyl IRNA s),nrherases

la
e\
ed

-\ 2-year-o1d boy with an ear infection was given amoxicillin, but it dld not clear up the problem. Switching to azithromycin successfuily eradicated the infection, and subsequent laboratory work indicated that the offending bacterium was resistant to amoxiclllin. Bacterial resistance to antibiotics is often due to whrch of the
[ollowrng?

(A) Coenzpre Q (B) Cholesterol (C) Dolichol (D) HMG-CoA (E) Ketone bodles

'.ttfl

e patient with type 2 diabetes has been prescribed

recombinant insulin to help control his disease. Three months after starting this regime, a blood test is done, which indicates that the patient is sti11 produclng endogenous insulin, in addition to the recombinant insulin the patient is taking. The blood resr has, ar irs basis, which of the following? (A) Posttranslational proteolytic processing (B) Posttranslational glycosyiation (C) Posttranslational modification of amino acid side
chains

iA) Altered ribosome structure

(B) Altered ceil wall (C) En4.matlc destruction of the antibiotic (D) Inability to transport the drug into the bactena (E) A mutation in RNA poll.merase

14.

E35L

of of

A young boy exhibits myopathy, encephalopathy, lactic acidosis, and strokelike episodes. Ail of his siblings have some aspects of the same s).rnptoms. The boy most Iikely has which type of mutation? (A) A defect in mRNA s),rrthesis (B) A defect ln mitochondrial IRNA production (C) A defect in mltochondrial rRNA production (D) A defect in cytoplasmlc IRNA (E) A defect ln cytoplasmic rRNA

(D) Posttranslationai acylation

(E) Posttranslational quaternary structure formation

"# o """""

Given the serine codons shown be1ow, what is the mlnimum number of tRNAs which is required to bind
to them? UCU, UCC, UCA, UCG, AGU, AGC

hospital laboratory made an error and mistyped a patients blood as AB, instead of B. When given type A blood, the patient had an adverse reaction. The major difference between individuals with AB and B type blood is due to which of the following? (A) The presence of a specific glycosyl rransferase (B) The presence of a specific acyl rransferase

(C) The presence of a speclflc peptidase (D) The lack ofdolichol pyrophosphate

(E) Increased levels of dolichol plrophosphate

{,tI

--:-,ir--.:

:
Introns are not found in mature mRNA (they are removed by spliclng in the nucleus), thus, intron-exon secondary

.stsIITR,S
! n-ir(:
Ite 5'cap.
The 5'cap ofmRNAis rec-

--E=:-=; yr- nririanon facrors (specifically eIF4E) to allow ::fi--B-rme assembly on the mRNA. The absence of a cap rr-Lruld not allow a translation initiation complex to form tsee the figure below for an oveMew of transiation initiation. Factor eIF4e is required for the mRNA binding to the smal1 ribosomal subunit through cap recognition).

structure would not be present. pseudouridine is found only in IRNA, not in marure mRNA. Th).,rnine, while found in IRNA by posttranscriptional processing, is not found in mRNA (uracil is placed into rhe mRNA when an adenine is in the template strand), and the poly-A tail, aL Lhe 3' end of the mRNA, adds stability ro the mRNA, but it does not play a role in translation initiation.

3'
Met
Small ribosomal subunit erF4E (A) + other eIFs

mRNA

Met
(oo)

_*---',-:r:-'-7/'\ --*---*'

\k

Met
ADP +
GTP Met-tRNAl/let
P;

Large ribosomal

subunit GDP +
eIFS
P;

EIF2-GTP.A.

(rerP

) \ Met-tRNAlvlet

::.:l:.. 1;r" answer is A: lnability

to initiate translation.

As

i,i$:i:: The answer is B: Reduced lysosomal enzyme

activity. The

muscle works, and AMP levels rise, the muscle wants to preserve its AfP for muscle contraction, and not to use it for new protein slrrthesls. The increase in AMP levels leads to the activation of AMP-activated protein kinase, which will phosphorylate and inactivate eIF4E (eukaryotic initiation factor 4E), which ls a necessary component in recognizing the 5' cap structure of the mRNA to allow ribosome assembly on the mRNA (see the figure in the answer to the previous question). The activation of the AMP-activated protein klnase does not alter

(mucolipidosis tlpe II), which is a defi.ciency in protein sorting, particularly of sending lysosomal en4rynes to the lysosome (a lysosomal storage disease). The I of l-cel1 disease stands for inclusion bodies. If the child develops these clinical and radiologrc slrrnptoms iater in life, one would consider rhe diagnosis of Hurler child has
I-ce11 disease

elongation or the termination of translation. It does not biock overall transcription, either of mRNA or rRNA (although it may lead to an lnhibition of ribosomal biogenesis as well as the transcrlption of certain specific
genes).

sl,ndrome (mucopolysaccharidosis). Lysosomal enzqes are tagged with mannose-6-phosphate (M6P) during posr translational modification. Enz).rynes containing M6P then bind to a M6P receptor, which transports the enz).,rnes to the lysosomes. Lacking such a signal, patients with I-cell disease secrete their lysosomal con[ents into the plasma and interstitial fluids. This leads to lysosomal dysfunction and cellular and tlssue destruction. The en4,,rne that is defective is shown on page 41. This disease is not peroxisomal, nor does it enhance protein secretion.

Protern

+1

7
O-Mannose

l-uop-NA"cr"

[/ I

phosphotransferase rdefective in I-cell disease)

Jruve
O-Mannose 6-phosphate-1 -NAcGlc

lrHro f N-acetytgtucosaminidase t\
O-Mannose 6-phosphate

fr ru-nce tc

is E: Inhibition of an elongation factor for translation. The child has diphthena. n-: :.- . caused by a bacteriurn (Corynebocterium diplthcriai which produces a toxin that leads to the inhr'br:----of eEF2 (eukaryotic elongatron factor 2), s'hich .s ' required for the movement of IRNA from the '.1 sr:e to the "P" site. The toxin catalyzes the ADP-ribcrsr lation (using NAD* as a substrate) of eEF2 to bnng about this inhibition. If one treats such a child n-rri
The answer nicotinamide (the reaction product resulting from the loss of ADP-ribose from NAD.), one can reverse anc block the ADP-ribosylation reaction catalyzed b;' the toxin. The toxin has no effect on protein kinase A. nor does it glycosylate a G protein. Diphtheria causes sore throat, fever, swollen nodes (bul1 neck), w'eakness, hoarseness, painful swallowing, and chi1ls. The hallmark of the disease is a thick, gray membrane co\'ering the pharynx.

-::reration of the mannose-6-phosphate slgnal on lysosomal pro:-ns

\\lthin the ER.

'1

The answer is D: Peptide bond formation. It is the large ribosomal RNA that catalyzes peptide bond formation, using peptides and amino acids in the "A" and "P" sites

on the ribosome. Destroying the secondary stl-Ltcture of this rRNA via the aberrant ribonuclease will limit
the abllity of the ribosome to create peptide bonds. The 1arge, ribosomal RNA molecule is not essential for the initiation, termination, elongatlon (movlng the ribosome along the mRNA after peptide bond formation has occurred), or IRNA activation and charging.

The answer is C: Chloramphenicol. Chloramphenicol blocks peptide bond formation in prokaryotic ribosomes (with no effect on eukaryotic ribosomes). This concept is the basrs of certain antibiotic therapy; the

The answer is B: M-A-D-S-G-M. The variant ce11 line will mischarge a tRNA"r" with glycine, but only the tRNA"r" that has, as its anticodon, IGC. IGC will recognize three

ln ribosome structure between eukaryotic and prokaryotic ce11s allow selective drug inhibrtion. The child has meningococcal meningitis, and chloramphenicol, despite its side effects of inhibiting mitochondrial protein synthesis, is a very effective agent for this disorder. Cycloheximide has the same effect as
differences chloramphenicol in eukaryotic cells but has no effect on prokaryotes. Rapamycln leads to the blockage of translation initiation, not peptide bond formatlon. Puromy-

codons: GCA, GCU, and GCC. Thus, when these codons are present in the mRNA, glycine w111 replace alanine. Thus, reading the RNA from the 5'end (as translation reads the

he
LSa

mRNA 5' to 3') provided in a senes of 3, we have AUG (which is methlonine), GCG (which is alanine-the anticodon for this codon is CGC; IGC will not recognize the GCG codon), GAC (whlch is aspartic acld), UCG (which is serine), GCU (which, in this cell line, is glycine and not alanine, since the lGC anticodon will recognize the GCU codon), and AUG (methlonine). The answer choices utilize the singleJetter codes to represent the amino acids.

cin is a chain terminator, stopping protein synthesis but not directly inhibiting peptide bond formatlon.
Rifampin inhibits prokaryotic mRNA s1,r-Lthesls and has no direct effect on translation. Rifampin might be used as prophylaxis for household contacts of meningococcal meningitis but is not as effective a treatment for the actual disease.

so-

lis;.

If

I
The answer is B:
ce1ls,

The answer is A: lnhibition

of mitochondrial

protein

I'|AD*. Diphtheria toxin, after entering

is cleaved by a protease to form an active enzyme,

synthesis. Ototoxicity (hearing loss) occurs with a subset of antibiotics because, in addition to affecting
prokaryotlc ribosomes, the drugs also have an effect

)ms

which, utilizing NAD* as a substrate, ADP-ribosylates

eEF2, thereby inhibiting protein translation. ATB FAD, acetyl-CoA, and UDP-glucose are not required for the ADP-ribosylation reaction. The final modified product, an argimne with an ADP-ribose attached, is shown below ADP-ribosylation: N of arg, gln; S of cys

rler
nes oslhen

on mitochondrial

ribosomes. Mitochondria contain their own DNA, RNA polymerase, and protein synthesizing apparaius (recal1 that it is thought that during evolution, bacteria invaded eukaryotic ce11s
itH,

sto
ce11

sma

tion

Adenlner

rt is
'oxi-

H\J
HO

f )

.o..

?*,-"-t-"-t_-o-?*:o..
HO
Answer 6

o J

il-3 --"r,-cH,-cH,J

OH

foy/-____;E_ \---{
OH

+2

.-:

,,:-

-:: :
inhibiting protein s).nrhesls. The drug does nor aflect
ribosome assembly The drug also has no effect on tran-

-,:-' ,--:::ei a s)mblotic relatlonship, with rhe bac.:r j -'.:niuaLl)- becoming the mitochondria), whtch ,: ..:1 similar io the prokaryotic apparatus. Thus, ..:iain drugs u.ill affect mitochondrlal proteln s)'n:lesrs, and the elfects seem [o be greatest on those rrtans that have high energy needs (such as neuronal tissue). Erythromycin does not affect mltochondrial RNA synthesis or DNA replicatton. it does not aflect the ear drum, nor does it increase neuronal signahng in the inner ear.
13

)cnption or RNA processing.

(: Ribosomal inactivation by covalent modification. Ricin, a toxin found in castor oi1 beans, specificall,v cleaves an N-glycosidic bond in rhe 2BS
The answer is IRNA of the large ribosomal subunit (an adenine base
is remor,ed, but the phosphodiester backbone remains

intact). The sequence of rhe IRNA rhar is altered

is

t0

to C(C. The most efficrent way to inserl a proline into the middle of lt, as proline cannor form an cr-helix due to the restriction of rotation of certain bond angles. Choice (A) has a leucine going to proline; all ol the
The answer is A:
CUC

lo

destro)r an o,-helix is

required for bindlng elongatlon factors dunng protein synthesis. As nbosomes become inactivated by ncin, protein synthesis in cells stops, leadlng to ce1l dearh. Ricjn does not inhiblt RNA polymerases or aminoerc;.l IRNA synlhetases. Ricin does noL initially aflect

ribosome assembly The LDro for ricin

bod1, u'eight.

is 30mg/kg

other substitutions to ala, leu to leu, ile ro va1, and to leu) would still allow cx-helix formation after the substitution took effect as Lhese are conser\rative substitutions.
(r,a1

va1

t4

tt

The answer is (: The drug binds to the 50S ribosomal subunit of bacteria and blocks translocation. Clarirhro-

mycin (an antlbiotic

in the macrolide lamily wlth

erythromycin and azithromycin) is specific lor rhe large ribosomal subunit of prokaryores (it will not bind to eukaryotlc ribosomes). When this drug binds to the large ribosomal subunit, translocation of the ribosome (movement along the mRNA) is blocked, which blocks overall protein synthesis. IRNA binding is not affecred by clarlthromycin, nor is there a blockage of the formation of an initlatlon complex. lt is the large subunrt (50S) that contains the peptidyl transferase acriviry, which is also not blocked by this agent.

The answer is (: Enzymatic destruction of the antibi. otic. Bactena that der.elop resistance to antibrotics usually. do so by contarning an enzymatic activity rhat destroys the structure of the antiblotic so that it cannot effectively inhibit its rarger within the cell Amoxicillin w'orks by, destroying the bacterial cell u,a11, by being incorporated into the growing cel1 wall, which ieads to a cessatlon of cell wal1 s,vnthesis. It does not alter ribosome structure. Mutations in RNA polymerase will not lead to resistance to drugs. Azithromycin was effective because the bacteria did not produce an enzyme that
destroyed the drug.

t5

The answer

is B: A defect in mitochondrial IRNA production. The boy has MELAS (Mitochondrial

myopath)., encephalopathy, lactic acidosls, and stroke),

12

The answer is D: The drug inhibits initiation

of protein

synthesis. Rapamycin inhibits the mammalian rarget of rapamycin (mTOR), which is a protein kinase. One ol the many rargers of mTOR is eIF4E brnding protein (eIF4E is a required initiation factor for protein spLthesis). When not phosphorylared, the bindlng protein binds tightly to eIF4E and prevenrs it from participatlng in the formation of the translational iniriation complex, thereby bLocking protein s1.r-rthesis. When phosphorylated at multipie locarions by mTOR, the bindlng protein fa11s off the initiatron factor and allows translational initiatron complexes to form. In the presence of rapamycin, mTOR has no kinase activity, and the binding protein remains bound to eIF,1E, thereby

a neurodegenerati\re disorder due to a mutation in mitochondrial IRNA, leading to defective protein synthesis wlthin the mttochondria. The component most often affected is complex 1 of the respirarory charn. The severity of the disease will be dependent on the extent of heteroplasmy (what percentage of the mitochondria codes for the altered IRNA). The
more mutant mitochondria present, the more severe
the symptoms.

l6 fhe answer is C: 3. A IRNA wlth the anricodon IGA will bind to UCU, UCC, and UCA (with wobbhng at
the thrrd position of the codon). A IRNA with the anticodon CGA will bind to UCG. And the third IRNA. with

Protein Synthesis
an anticodon of ICU, can base-palr with both AGU and AGC. Recall that l wil1 base-pair with A, C, or U in the wobble positlon. These codon*anticodon interactions are shown below.

43

space around the tissues. Thrs ieads to rhe protruding abdomen seen in these stan-ing individuals. The prob-

A.

Codons for serine

s'

c u-3' -uUCC
UCA UCG AGU AGC

lem is not related to reduced muscle protein synthesis (which does occur. but does not alfect the osmotic strength of the blood), or reduced intesrinal protein synthesis, for the same reason as the muscle. Both protein hydrolysis and tnglr-ceride hr-c1ro1y,srs require
water; water is not produced ri hen these molecules are

broken down.

18

B.

Base pairing of serine codons with anticodons

3' on mFtNA -u ? !i : 3',-A G : _, Anticodon t-5' o"inr.rn+r

5
s', rr r -A Y c-

codon

Codon on mRNA Anticodon on tBNA #2 Codon on mRNA Anticodon on IRNA #3

-U 5 _U
3'

3',

,:, i
C

-, "

9- 3 I " -. "

The answer is C: Dolichol. The biosrnthesrs ol both coenzyme Q and dolichol is dependent on r.LrDrin.units, specifically isopentenyiplrophosphate uhich rs derived from mevalonrc acid in the de novo p.1rh\\-:i\ of cholesterol biosynthesis. Lovastatin inhibirs H\lGCoA reductase, which reduces mevalona[e producrir]r-t This can have, then, the unintended consequenre it ,r reduction in dolichol production, thereby leading to underglycosylation of processed proteins. A reduction of coenzyme Q synthesis can lead to muscie weakness (a slde effect ol the statin class ol drugs), bur coenzlme Q is not involved in protein glycosylation. Cholesrerol, HMG-CoA, and ketone bodies are not required for protein glycosylatlon as well. The srructure of dolichol, in r,vhich the isoprene building block is highlighted, is shown below

-A

ooHIcH.l"r"
o - B -o - [ - orrrll-

oocH.ll" "rr-.r,-

J-

"r,-.]

"",

-",

J-"r. -.r, | "rr- "*=

l7

The answer is B: Lack

of Iiver protein synthesis. The

boy is displayrng kwashiorkor due to a caloriedeficient diet 1ow in protein. Since the boy is taking in less protein than he needs, he is becoming deficient in the essentral amrno acids. Thus, to syntheslze new proteins, existing proteins need to be
t

l9

The answer is A: Posttranslational proteolytic processing. As shown on paEe 44. insulin is s;,mthesized as a prepro-

I
l

degraded such that a pool of essential amino acids is available for the new protein synthesis. This protein degradation occurs in the muscles, leaving the boy with very thin arms and legs. The 1iver, despite the increased muscie protein turnover, is sti11 deficient rn essential amino acids and reduces lts level of protern synthesis, including those proteins normally found in the blood, such as serum albumin. The reduced protein content in the blood reduces the osmotic strength of the blood such that when the blood flows through the body, the osmotic strength of the tissues is hlgher, and fluid leaves the blood and enters the interstirial

insulin. The pre sequence is the signal sequence, which is cleaved when the protein enters the endoplasmic reticulum. Proinsulin contains the C-peptide, which rs removed to form mature lnsulin. and dlsulfide bonds that hold the A and B chains together. The modification is thus proteolytic processing, and not glycosylation, modification of srde chains, acylation, or altered quaternary struciure. The test is a C-peptide level. If a person is producing insulin, a C-peptide 1eve1 can shou, this. This is also a way to see if a person has type 1 dlabetes mellitus (no C-peptide, slnce they cannoi produce endogenous insulin) or type 2 (normal or high C-peptide levels, slnce this disease is a reduced response to normally produced insulin).

Ti
44
Chapter
5

A chain

b a

B==-.-,
(*
Bif,iLxi''"

.={!,9
IE

''\.4
sulin to mature lnsulin.

il

f,

ifr

.--+
I

Golgi apparatus

NHg+

s-s{l
)

"oo.,
Signal sequence

,/,,
C-peptide

Preproinsulin

Proinsulin

Answer 19: panel A rndicares the amino acid sequence of marure insulin while panel B indicates the steps involved in converting preproin-

20

The answer is A: The plesence of a spetific glycosyl trans'

ferase. As shown below, type A and B blood differ by the presence of one sugar on glycosylated proteins. Individuals with tlpe A blood add one tlpe of sugar (N-acetylgalactosamlne), tvhile individuals with type B
blood add
a

in the specificlty of a glycosyl transferase that recognizes the base carbohydrate structure. The dlfferences in blood group antigens are not due to acylation or proteolytic
processing on the ce11 surface. As these carbohydrates are Olinked, dohchoi (whlch is required for N-linked glycosylation) is not required for their rynthesis.

drfferent sugar (galactose) due to dilferences

Blood type
Type O

G)"@>"8
H substance

@>"8
Type B

(;lx"<*^>"8 -.\_--]/ \-

G)
-Ol

Structures of the blood group antlgens.

l
Chapter G

Expressian
tlils chapter, the questions will cov*tt ttartotts :rtp.,.|r, o{ Xyne re gulation, focwsing on the ways ut y'hich both prolwtryotes and eutzalyarcs re gttlite
In"
lt o

Regulari*n of Gene
sickle ce11 disease. His sibling shows no syrnproms rrt the disease, although genetlc iests showed iro*or1-gor_ ity for the HbS gene. An analysis of his red blood ce1-k is 1ike1y to show which of the following?

th tu anscrip,yion and ty ansl ation.

QUESTT0NS
Select the single best answer.

i
tlzes

iood
ir tic
-ales Lked

one partial diploid, expression of the lac operon is still constitutive (syrrthesrs of the genes is observed even in the absence of an inducer). A likely explanation for this resuk is which of the following?

the genes of the 1ac operon (constitutive sl,nthesis). The scientist creares partial diplolds of rhe regulatory elements of the 1ac operon in these mutants of E. coh. in

Wh11e studying the 1ac operon in bacteria, a sclenrisr isolates mutants of Escherichiqcoli, which always express

(A) Reduced alpha chain s1-nthesis (B) Reduced sickle chaln synthesis (C) increased gamma chain sl,nthesis (D) Increased zeta chain sprthesis (E) Increased delta chain strrthesis

A healthy reenage girl has .orn. ,o her pediatrician for a presports physical. Results of hemoglobin elecrrophore_ come about factors
sis indicated an elevarion of fetal hemogiobin. This can
r.-ia

(A) Overall increased expression of all rranscription (B) Overall reduced expression of a1l transcription
factors

which of the following mechanisms?

(A) There is a muration in cis wlth the operon (B) There is a murarion in rrans with the operon (C) Inducer can no longer bind ro the repressor (D) Inducer binds too rightly ro rhe repressor (E) The transaoivatton domain ol the repressor
mutated

(C) Deletions in the locus control reglon of the B_globin

gene cluster

(D) Deletions in the locus conrrol region of the o_globin

is gene cluster

(E) Inappropriate looping

OCCUT

2 An African Amerlcan pattent

of chromosomal DNA, a1low_ ing transcriptron of prel-iously inaccessible genes to

has dlsplayed

vaso_

rvhich of the following?

occlusive episodes for most of his life. The incidents are more prevalent under conditions in which blood oxygen levels are low, such as dunng exercise or taking t.ipi,o locations at high ahitudes. The patient has been placed on hydroxl-r-rrea. The rationale behind this treatment is
To prevenr vaso-occlusive episocles through hydroxy_

5 ,l

(A)

urea induced protein degradation

(B) To reduce syrrthesis of a defective protein (C) To induce sprthesis of a functional protein (D) To enhance or)-gen levels in the blood (E) fo acrivare rhe enzyme that produces 2,3_bisphos_
phoglycerate

a srudy wrrh mice exhibiting hypercholesterolemia, cholesrerol was affixed to double-siranded RNA, rvhich targered the dsRNA ro enrer ce11s through cholesrerol diflusion through the plasma membranel The dsRNA was targeted ro bind to mRNA that encoded the aoolj_ poprotein B gene and resulted in a iowering of circllar_ ing cholesterol levels. This result occurs due to which ol the follor,r.ing?

(A) Inhibition of apolipoprorein B rranscription (B) Inhibltlon of apolipoprotein B rranslarion (C) Inhibition of apolipoprorein B folding (D) Enhanced degradation of apolipoproteln B (E) nNe editing of the apolipoprotein B mRNA

A hematologist is studying an African American family as one of the children was recently diagnosed wlth

6
45

A woman wirh a BMI of 16.5 visirs her family phr.sicr...:: because she always feels tired. The historr..indicates ::,:-

r : :l- ::S a\ eI 3 h/da),', - -- .,. --:^ indicaies thal : : '. la-'l\\- nolmal Under .,::- : --.1:,1t\- lO malntain hef
:-- ::r,11 results,
,-

to

in part, from

.o$ing proteins?

Theoretically., a disease could result from an increased expression ofa particular gene. This can occur in eukaryotes through a single-nucleotide mutarion in a promoterproxrmal element. Thrs is best explained by r,vhich one ol the follolving?

fA) \ltrrc elficienr 'plicc sire recognirion (B) lncreased opportunrty for hydrogen bonding ro a
transacting lactor

:::.

subunit 56 hormone receptor

(C) Beneficial amino acid replacement derived from the

missense mulatioll

(D) lncreased amount of sigma factor binding

--r:19-slandlng patient of 1,6n.t has der,eloped multi:,e tumor tvpes during his lile (,11 years old). You har.e dragnosed him as har.ing a specific s)mdrome involving p53. The multiple cancers that result from this s),ndrome are pnmari11, due to r,vhich of the follolving initial drrect efiects of the inheriLed mutation?
---

(E) Reduced energy need to melt rhe DNA helix at this

position

il

\)

lmprrred gene transcription

A human genetic condition in whrch too much ol a gene is routinely expressed has been mapped to a locus on a dillerent chromosome from where the gene in question is located. Which one of the following is a porential

lB\

tnhrnced gcnc transcriprion

explanation lor the condition?

(C) Impaired protein sl.nthesis (D) Enhanced protein s),nthesis (E) Altered chromosomal strltcture

(A) The activated gene has a TAIA box murarion (B) The locus control region for the gene is delered
(C) (D)

gene encoding a iranscriptional repressor has

been mutated

A patient has been prescribecl methorrexate for chemotherap): After an initial success in reducing tumor
growth, the lumor resumes its raprd gro\\rth. One potential mechanism lor this is wl-rich of the follorving? (A) Amphf,cation of the dihy.drololate reductase (DHFR)
gene
1Z

transcriptional actl\,ator sustained a missense mutation, u,hrch reduces rts alfinity for DNA
a

(E) A variant promoter region is lormed owing to

splice site mutation

A patrent is taking

(B) Amplification ol the gene lor degradrng merhorrexare (C) Reduced transcription ol the gene allolvrng merho, trexale enlr).into the cell (D) lncreased transcription of the gene a1low.rng methoirexate efflux from Lhe
ce11

cy closporin A afrer receiving a kldney transplant. Cyclosporin A protects agarnst organ rejec-

tlon bv r,vhich ol the lollowing mechanisms?

(E) An inacti\.ating mutation in the gene for DHFR

16-year-old girl has been loslng weight and feeling lethargic over the past 4 months and is raken to the physlcian by her parents Durlng the histor1,, the parents expressed concern that their daughter had seemed to eat yery little during the dall a claim denred by the patient. Laboratory results indicated an iron deficiency and a microcytic anemia. The cel1s of the patient har.e adapted to the iron deficiency rn which one of ihe

(A) Blocking translalion of cytokine genes (B) Actir,aLing transcription of cytokine receptors (C) Stimuiatrng the phosphorylation of transcription
lactors

(D) Blocking the dephosphorylation of specific rranscflption factors

(E) Stimulating translation of cytokine t3

genes

following ways?

(A) Increased transcriptron of ferritln mRNA (B) Reduced transcnption of the translerrin receptor
mRNA

patient has asthma, but has become resistant to glucocortrcoid inhalation. A potential mechanrsm for this resistance is w'hich of the following? (A) lnabilrty of glucocorticotds to enter targer ce1ls (B) lnabihty to induce histone acetylation (C) Reduction of leveis of transactir.ating faciors in rhe
nucleus
kinases

(D) Cytokine lnduction of protern

(C) Increased translatron of the ferritin mRNA (D) lncreased translatlon of the transferrin receptor
mRNA

(E)

lncreased dimerization of the glucocorticoid receptor

(E) Increased degradation of the transferrin receptor

mRNA

14 lnductlon of
decrease

certain transcriptron factors leads

to

in the expression of certain genes. This occurs

through r,vhich ol the lollorving mechanisms?

Regulation of Gene Exprls.

ed
erne

--:' +7

(A) Decreasing the rate of RNA polymerase catalyzed

(A)

Lossofionicin|eractionsbetu.eenthetral:'::'
factor and DNA

(B)

(C)

)a

(D) (E)

phosphodlester bond formatlon lnducing the s1,r'Lthesis of a protein that posttranscriptionally edits mRNA such that translatron initiation is blocked By decreasing the rate of RNA po1y.6.trt. binding to the promoter Through stimi-llation ol proteins w'ith HAf activit)' lncreasing enhancer bindlng to DNA

in hydrogen bonding betr.veen tra.s-::' lactor and DNA tion (c) Decrease in hydrogen bonding betr,veen the t:,-.: scription factor and DNA (D) lncrease in ionrc interactions between the transci----tion factors and DNA
(B) lncrease (E)

lnability of
DNA

Lhe transcrlption lactor

to bind to thr

l5
his

Shown below is a partlal map of the promoter region, and promoter-proximal region, lor the y-globin gene. The overlapplng binding sites for transcription factors allow for which ol rhe followrng to occur?

The next two questions refer to the following situation. In order to better analyze the promoter region of a particular gene, this cloned region of the gene was placed in front of a
reporter gene and the resultant vectors placed in eukaryotic cells to measure the expression of the reporter, using various deleted constructs. The results obtained were as follows:

lne on
.e

s-

tial

lonrnllcnrnl
-175

@ @ @ @ l*],'';"'.'o''on cAAr cAAr rArA I

site

-l 15 -85
tl-Le

-30

Construct ABCDEI-1oo

Reporter level (relative)

Aspecrs of

y-globin gene promoter.

(A) The ability to modulate the binding of positrve, or

has
nse

negative, transactlng factors to the DNA

(B) The

abillty to reduce the risk of losing transcrip-

r*
r-----|

oa

tional control r.ia mutation in this reglon (c) Promoting looping of this DNA region (D) Providing a Larget for interfering RNAs (E) Providing ribosome binding sites for translation ini-

150

tiation
nev
i

10

ec-

lron

A hypothetical patient was suffering from excessrve free iron in the blood, yet a ce11u1ar analysrs indicated iow intracellular levels ofiron, despite high intracellular 1evels of ferritin, and normal transferrin 1evels in the blood. The disease was shown to be caused by a single base change rn the DNA that led to a dysfunctlonal protein. The mutation is hkely to be ln which of the following
proteins?

42oo -

(A) Tiansferrin

LtO lor

(B) Translerrin receptor (C) Tianscobalamln (D) Iron response element bindrng proteln (E) Ceruloplasmin

lndrcate which region (as deslgnated by the letters A, B. C, D, and E) binds an inhibitory transcription factor.

r9

Referring to the figure above, which region binds stimu-

Iatory transcription factors?

A woman developed the following syrnptoms after taking cer[ain drugs such as barbiturales. The s)rmptoms included severe pain in the abdomen, hallucinations.

t7 A cell is producrng a certain transcription factor that

r Lhe

)ptor

Loa
]CUTS

coniains a single point mutation, such that N is converted [o V Circular dichroism experiments show that thls altered factor has the same secondary structure as the nonmutated factor. Under normai conditions, serum stimulation of quiescent cultures shows strong induction of fi.ve genes. When quiescent cells harboring the mutant transcriptlon factor are exposed to serum, the level of expression of those flve genes stays at basal leveis (there is no increase in mRNA production). This finding is most likely due to which of the followrng?

drsorientation, and a reddish tint to the urlne. These symptoms appeared due to the induction of genes involved in which of the following pathways?

tA) Cytochrome s)mthesis (s) Cl tochrome degradat ion

(C) Ubiquinone brosl.nthesis t D) Ubiquinone degradation (E) Dolichoi s)Trthesis

48
1

Chaprer 6 are adjacent to the operator. Thus, if a partial dlploid contains a normal operator region on the extrachromosomal region of DNA, that operator region cannot regulate the operon on the chromosomal DNA. Thus. the constltutlve mutant that was not rescued in a partial diploid ls mosr likely an o. muration. If the inducer could no longer bind to the repressor, then no expression would occur, as the repressor would not leave the operator region. In addition, if this were the case, then adding normal repressor to the cel1 (via the parrial diploid) should a1low expression of the operon. Similarly, ii inducer bound too tightly to the repressor, the rntroduction of normal repressor should reyerse the effects of the mutated repressor. The lac repressor does not conlain a transactivation domain.

ANSWERS
The answer is A: There is a mutation in cis with the operon. Constitutrve s1.r-rthesls can occur by either of the tu-o mechanrsms. The first is an inability to s;mthesi:e lac repre ssor; the second is to have a mutation in the operaror region that renders repressor binding impossible (an o'mutarion; see the figure below). An inability to s\nthestze 1ac repressor can be repaired in trans; if rhe partial diploid contains a functional lac repressor gene, functional protein will be sprthesized from the gene. which can brnd to the chromosomal operator region, and regulate 1ac gene expression. If, howeyer, an o'mutation occurred, the operator region is in cis with the operon and can only regulate regions of DNA that

lnducers
Promoter Operator Structural genes

ABC
No transcription occurs No proteins are produced

(active)

Bepressor
tnducer

I I

Bepressor

ry/

o--i
I I

(inactive)

RNA polymerase

polycistronic@ mRNAltl
Protein Protein

tir

ABC

Protein

An overview of the regulation of the 1ac operon (an inducible operon). In the absence of an inducer, the repressor binds to the operaror, preventing the bindlng of DNA poll. nerase. When the inducer is present, the lnducer blnds ro the repressor, inactivatlng rt. The inactlve repressor no longer blnds to the operator. Therefore, RNA poll.merase can brnd to the promoter reglon. If there is a mutation in the operator reglon such that the repressor can no longer bind (an o' muration), constitutive expression of the operon (expressron in the absence of inducer) will
resuit.

ol a functional protein. The patlent has sickle cell anemia, and hydroxyurea treatment is designed to acrivate rranThe ansurer is C: To induce synthesis

scription of the y-globin chain, which is normally only expressed during development (feta1 hemoglobin). When expressed, the y-globin gene will form functional hemoglobin tetramers wlth the cx-globin chains, thereby reducing the effects of the mutated B-globin chain. Hydroxyurea does nor bind to hemoglobin

it; it does not reduce the synthesis of the B-chains, nor does it aher oxygen levels in the blood or 2,3-bisphosphoglycerate levels in the erythrocyte. The y-chain is normally rurned off at birth as part of the hemoglobin-switching parhway (see the figure on page 49). The challenge ro scienrisrs ar this time ls to understand how to reactivate y-chain synthesis in patienrs with both sickle cell disease and
and denature
B-thalassemias.

la1 diploid

Reguiatlon of Gene Express:c::

Chromosome 16

+9

extrachro.on cannot NA. Thus. I in a par-

-. HS4o
Chromosome

u,

0,

re inducer
:1O

11

expres-

r leave the case, then

-.lqFuGvAvDB
Embryo: (2er= Gowel1
(zYz = Portland

artial dipimilarly, if introducects of the

0'2t2= Gower 2
Fetus: q.2y2 = HbF

contain

Adult: o2y2 = HbF

0252 = 42

dz9t = A

'6 o

50

insrgr ):

c Changes

r@-11::::.:it. Panel A indicates the chromosomal

in globin chain expression dunng

loca_

E,
o o

6'"

v:rl.e !::-arn expression begins to increase.

at which n*q - : ::-. elopment a particuiar ctraln rs expressed. Note ma ::-:'-:-j y-chain expression is reduced signiflcantly

t^= :r -::: giobin

chains while panel B indicates

.-0

18
Prenatal age (wk)

30

Birth

postnatal age (wk)

answer is G: lncreased gamma-chain synthesis. The -=-:iing has, in additlon to sickle cel1 disease, hereditary

f*

-rsisience

of fetal

express rhe y_globin chain throughout their 1ife, =:rd ir can be at high levels. Since this cltld is express_
.1S

:?FH

hemoglobin (HpFH). Individuals wirh

in all transcrlption factor expression in the celi. While inappropriare looping may help to lead to y_giobin gene expression, the looping needs ro be modulaf,d by [arr_ scription factors for gene expression to occur.

rtor, preepressor rr reglon cer) u-ill

the HbS prorein and the y_prote1n, some nor_ ::-ai letal hemoglobin can be formed-in this child, with :tduced levels of HbS formed. This reduces the 1evel of ickling and aliows oxygen delivery ro rhe tissues. Thus, -re HPFH protects againsr the effects of homozygous -{bS expression, and the sibling shows few, if any, ffip_ :lms of hls HbS mutations. Alterations in the expression -.i rhe 6-chain have nor been observed. Reduced

both

$ "' fne ansurer is B: lnhibition of apolipoprotein B translation. The cholesterol tag on th!-isnNA allowed

tre \tay to reduce the syrthesis of Hbs chain. increased I-trrthesis has also not been observed.

s-.nrhesis would lead to an anemia, and there is no eilec_

c,_chain

Lesis

of

in

the
as

eryrh-

rirth

;ee the

:egon of the B-giobin gene cluster (since fetal

Eiobin is

This can come about by deletions on the locus control

T.."rYe1 of the B-globin gene cluster. The girl is expresiing HPFH (hereditary persisrence of feul hemogiobln).

i-s-C: Deletions

in the tocus controt region

at this

n slTr;e and

uanscription factors would not lead to increased tran_ scnption of the y-chains, nor would a general increase

orB, mutations n the locus control region ol the o_gene cluster will rot affect fetal hemoglobin synthesis). "A general loss

arf,

hemo_

and adult hemogiobin is

lead to either the destruction of the mRt tA or the blockage of translarion of the mRNA. in either event, rhere wr]t Ue a reduction in apoB translation such that cells can no longer produce apoB100 or apoB4g. The dsRNA does not affect the transcription of the apoB gene, nor does it interfere with apoB folding once it becJmes rranscribed and translared. The dsRNA does not affect the rumo\.er of the apoB protein, nor does it edit the apoB mRirrA (other sysrems in the cell will do that). This situation was first reported in SoutschekJ et al. Nature. 200,1 Nov 7 I ;432(7 0 t4) : tT 3 -17 8.

ce1ls to take up the dsRNA, which was processed byintracellular ribonucleases to make a specific silenc_ ing RNA lor the apolipoprotein B mRNA. Binding of the processed dsRNA ro the apoB mRNA wiil

of

i*i;: ,,t"

ansurer c: GREB. CREB (cyclic AMp respon:e .is element binding protein) is a transcription factor that ts activated by protein kinase A and thai regulares, in pan. the expression of phosphoenolpl,ruvate"carboq.kirLase

50

Chapter 6
(PEPCK), a necessary protein for gluconeogenesis. Since

the patient is anorexic, her blood glucose levels

are

being maintained primarily by gluconeogenesis, and the enzFnes lor that pathway need ro be upregulated. The release of giucagon and epinephrine, both of whlch would be elevated in this patient, leads to the activation of protein klnase A and an increase in gene transcription for those genes regulated by CREB. Under these conditions, prorein slnthesis wlll be hmited, so factors necessary lor protein s)nthesis would not be generally acrivared (elF4, eEF2, and ribosomal protein 56). Nelther glucagon nor epinephrine works through steroid hormone receptors (they both utilize serpentine receptors on the ce11 membrane).

result of secondary mutations, but not directly from the mutations in p53) or to alterations rn protein sy:ithesis p53 mutations also do not alter chromosome structure.

The answer

is A: Amplification of the dihydrofolate

reductase (DHFR) gene. Methotrexate resistance most often occurs due to amplification of the gene for DHFR, the target for methotrexate treatment. Through overpro-

duction of DHFR, there is sufficient enz)ryne available to overcome the effects of the drug given to the parient. Resistance does not come about by altering the rate of entry ol the drug inro rhe ce11, by inactivating DHFR, or by inducing an enzyme that can degrade methorrexate.

The answer is A: lmpaired gene transcription. Individuals with Li-Fraumeni sl.rLdrome inherit a mutated copy ofp53, the product of the TP53 gene (on chromosome 17). p53 is a transcripiion facror whose major job is to monitor the health ol DNA, if DNA alterarions are found, p53, acting as a transcription facror, u-i1i iniriate new gene transcnption to arrest the cel1 cycle until the DNA damage is repaired and to also induce genes necessary for DNA repair. If the DNA cannor be repaired, p53 wili initiate gene rranscnption leading to cellular death (apoptosis). In the absence ofp53 acriviry, damaged DNA will be replicated, which increases rhe probabrlity of errors, eventually causing a mutation that leads to a cancer. Thus, the inttial inacrivatrng evenr ls impaired gene transcription by p53, whrch is the trlgger for al1 other events that fo1low Mutations in p53 do not 1ead, directly, to enhanced gene transcription (this may occur as a
Transferrin receptor synthesis
mRNA mRNA

of the transferSrnce the parient has an iron deficiency, leading ro a microcytic anemia, cells will upregulate their mechanism for acquiring iron, which is through the transferrin recepror. Ferritin is the iron storage protein within cells, and if intracellular iron 1eve1s are low, there ls no need to upregulate the s1,r-Lthesis of fernrin (its qmthesis is actually downregulated under these conditions) The iron travels in the circulation bound to transferrin, so increasing the number of transferrin receptors on the ce1l surface will enable a more efficlent transport of iron and transferrin into the cells. The regulation of transferrin receptor sylthesis is at the 1evel of translation, as is the regulation of ferritin synthesis. Thus, cells under these conditions will increase their translation of the transfernn receptor mRNA. This translational regulation is shown below.

The answer is D: lncreased translation

rin receptor mRNA.

Ferritin synthesis

IRE-BP
5',

IRE

AnJ

No translation

QIron
lron

CI

/|RE-BP

( ) s rz{hnnnn A,fi11, o"r'

1r,"n",,,,"n
JTranslatior
Ferritin

e/lRE

TRibosome

IY

Answer 9: Tianslational regulation of ferritin and translerrin receptor slnthesis via the iron-response eiement binding proteln (IRE-Bp) When
the IRE-BP binds iron, it w111 dlssociate from the mRNA. When this occurs, the transferrin recepior mRNA is degradeJ(since there are adequate tron lel'els within the cell, there ls no need to transfer more iron lnto the celi) and the ferririn mRNA is translated to produce more of this rronstorage molecule.

I-

Regulation of Gene Expression

5I

The answer is B: lncreased opportunity for hydrogen bonding to a transacting factor. A single nucleotide

.--:stitution in a promoter-proximal regron has the

ite
--.:.:bility of increasing hydrogen bonding to a transact= ,actor, thereby increasing the interaction betu,een --: factor and DNA and enhancing recruitment of .-t polymerase to the promoter. Slnce the mutation .:r the promoter region, it is not involr.ed in splicing -rch occurs at exon-intron borders), it does not code : ...n amino acid (since exons are not in the promoter , i...n), it is not related to sigma (which is prokan'otic :.:rfic), and the DNA helix does not have to be signifi-. :-r melted in this area to allow transcription factor : trlng.
The answer is : A gene encoding a

Drug treatment does not dlrectly affect the translation ol cytokine genes (it is indirect because the mRNA lor these genes is never made), nor does it lead to the actr\-rtiLrn ol transcrlptlon of cytokrne receptors. Cyclosporin does not lead to the phosphorylation of transcription factors. nor does it stimulate translation of cytokine genes.

--

r3

tion.

fhe answer is B: Inability to induce histone acetyla. Glucocorticoids bind to a cytoplasmic receptor and translocate to the nucleus,

:
ter-

transcriptional repres-

sor has been

: -

-..ssume

mutated. For the sake ol this ansu,er, 1et that the overexpressed gene (named A) is

. --..-ed

on chromosome X and the mutated gene (named

-.locatedonchromosomeY Thegeneonchromosome a transcriptional repressor that binds to --: :rromoter regron ol the gene A on chromosome X. ,'. ::r rhe repressor is expressed, gene A transcription . ::iuced. The protein product of gene B is acting rn .,,::s rn regulatrng gene A expression. A mutaiion that :,,,:i\ates the protein product of gene B, then, rvould . --rable to repress gene A transcnption, and lead to ::arpression of gene A. 1l the promoter for gene A -,: l TAIA box mutation, one would expect re duced ,,::':ssion (due to an inability of the basal transcnp. :: complex to bind), rather than enhanced expres. -:, *i161ln1ly, if the locus control region of the gene -. ::leted, then reduced expression for gene A would

. producing

r,vhere they bind to glucocorticoid response elements on the DNA, leading to a complex of proteins at this site. A number of transactlvating factors recrulted to the DNA contain histone acetyl transferase (HAf) activit),, which leads to further unwinding of the DNA from the nucleosome, enabling gene transcription. Patients r,vho become reslstant to glucocorticoid ireatment show a reduction in histone acetylation in response to the drug for a variety ol reasons. The resistance does not appear to be due to rnabrlity of the drug to enter target cells. Once inside the cel1, the drug r,vr11 bind to the receptor, and dimerizarion of the receptor is normal. In some resistant individua1s, the dimerized receptor has trouble translocating to the nucleus, rvhich leads to the reduction in HAf activity ln other patients, however, translocatron is normal,

and the reduction rn HAT actlvity appears to be due to an inability to recruit transacti\.ating factors to the complex, which contain the HAf activitl The levels of transactivating factors are normal, but it has been
hypothesized that the receptor is phosphoryiated under resrstance condrtions, leading to an inability to attract the transactivating factors.

.:rpected,notenhancedexpression.Ifatranscrip
-:,.i1 activator (gene B) suffered a missense mutation :, reduced its affinrty lor DNA, there r,vould be less -.-:,slription of gene A, not enhanced expresslon. And, -,..i. promoter regions do not undergo splicing.
TLe answer is D: Blocking the dephosphorylation of spe-

fi

cffic transcription

factors. Cyclosporin A binds to the .:rn cyclophihn in immunocompetent lymphocytes, --, -- :ris protein complex leads to the lnactivation ol ca1- -. .::in. Calcineurin. in response to increases in c; Lo,---.::.ric calcium, will activate its phosphatase activiiy and

The answer is C: By decreasing the rate of RNA polymerase binding to the promoter. Tianscriptron factors can be either positive or negative acting. In either event, to exert an effect on transcription, the factor must brnd to the DNA and then either promote RNA polymerase binding to DNA (in which case, lt is a positlr.e-acting factor) or.inhibrt RNA polymerase bindlng to DNA (in which case it is a negative-acting factor). Negative acting factors can do so by blocking the bindlng of positively acting factors to the DNA or by blocking necessary transactivating factors from binding to other factors already bound to the DNA. In either event, the net result is a reductlon in the rate aL which RNA polymerase binds

-:

:1!rsphorylate cytoplasmic nuclear factor of activated ,-.-.:rphocytes (NF-AT), a transcription factor. When
'.,,-,i inieract

to the promoter region ro iniriare transcription. Once

initiated, the rate of phosphodiester bond formalion is constant. RNA edrtrng is a rare event, and is not used to regulate gene transcription. If HAT activlty were stlmulated, gene transcnptlon would be increased, as the acetylated histones would have a reduced affinity for DNA, and rt would be easier for RNA polymerase to bind to the promoter regron. lncreasing enhancer bindlng to DNA wouid also increase the rate at which RNA pol1,n-rerase would brnd to the promoter, as enhancers

-.:::osphorylated NF-Af will translocate to the nucleus,

with nuclear factors and bind to DNA,

-,:ting new gene transcription. When calcineurin is -., -.i-ated lta cyclosporin-cyclophilin binding, NF-AT
-i:.:-Lrt translocate to the nucleus, and cytokine rynthesis --e cell (second messengers) rs compromised. Immu- - ---rnpetent cells have very 1ow- levels of calcineurin,

--,,r make them susceptible to cyclosporin treatment.

D*

--ir,l :l l

j -:::1- :he association of positive-acting transactiva--- :::iors, which would promote RNA po1;''rnerase
-:-:-ilg to the promoter
and transcription.

l6

It

The answer is A: The ability to modulate the binding of positive, or negative' transacting factors to the Dl'lA' The recognition of overlapping sequences on DNA by different factors allows either positive-acting or negative-acting effects, depending on which transcription factor is present in hlgher concentration. The y-globin gene is turned

The answer is D: lron response element binding protein. As shown in the figure below, the lronresponse element binding protein (iRE-BP) binds to loops in RNA (RNA secondary structure), and ln the case of ferrlti.n mRNA, the IRE-BP blocks translation when rron levels are low. In the case of the transferrln
receptor mRNA, the IRE-BP stabilizes the mRNA when iron levels are low, allowing efflcient translation of the

off after birth, and this has to do, in part, with factors bound to the promoter. When stage selector protein (SSP) is bound, y-globin rynthesis is favored, but when SPI blnds to thls area instead, y-globin s)'nthesis is repressed' A similar story occurs at the CAAf sites; when CP1 is bound, y-globin rynthesis is favored, but when CAAf displacement protein binds, CP1 can no longer bind, and gene transcription is inhibited. Thus, y-globin expression is regulated by the concentrations of both positive-acting
and negative-acting factors available in the cell The overIapping blnding sites were not designed for redundancy in case of mutation, to provide ribosome binding sites (the ribosomes bind to RNA produced from exons, not from the promoter reglon), or to provide a target for interfering RNAs (the siRNAs are targeted to mRNA) The multiple

binding sites, by themselves, do not promote looping of DNA, but once transcription factors have bound to the DNA, looping may occur as the transactivating factors bind to the proteins bound to DNA

mRNA such that transferri.n receptors can be sprthesized and placed in the membrane. lf a patient had a muiated IRE-BB whrch could no longer bind to its target sequence ln mRNA, then the transferrin receptor mRNA would be unstable and degraded, and ce11s would not be able to take up lron from the circulation, leading to higher than normal iron levels in the blood yet stllI would result ln ce11u1ar depletion of iron. However, the lack of IRE-BP binding would al1ow ferrltin translation and slnthesis, leading to high leveis of ferritin inslde the ce11 desplte the low Ievels of intracellular iron. Ceruloplasmln is involved in copper transport, not iron A mutation in transferrin would lead to lower circulating iron 1eve1s in the blood as the iron carrier would be mutated. Mutations in the transferrin recep[or would not lead to high intracellular 1eve1s of ferritin (lntracellular iron leveis would be low, the IRE-BP would remain bound to the ferdtin mRNA, and translation would be

blocked). Mutations

in transcobalamin would affect not iron. vitamin B,, transPort .

Transferrin recePtor sYnthesis

mRNA
An 3'

Ferritin synthesis

IRE_BP ../

IRE

Ribosome

ryl\"'
I I
Transferrin

mRNA 3 s',\,AllN\,/\,/1,/\,/\ \ow

iron

r\n

r\,/An

3'

receptor

t
lron

No translation

| -6t,on
r,on

Q lron

f,--*c

,lRE-BP

C)

,-IRE-BP

5'\.^n^n^^r.lh,,"l["ro"t'
I mnxn
I
no translation Desraded

i'?o

E desraded \-/
mnr'rn

5,

rnl ^rn rA\1

element blnding protein Answer 16: T?anslational regulation of lerritin and rransferrin receptor sy,nthesis lra [he rron-response (since there are the mRNA. when this occurs, the transferrin receptor mRNA is degraded when the IRE-Bp binds lronlt will dissociate from is translated to produce more and the ferritin mRNA adequare iron leve1s within the ce11, there is no need to transfer more iron into the ce11)
of this iron-storage mo1ecu1e.

(]RE-BP)'

Regulation of Gene Expression

The answer is C: Derrease in hydrogen bonding between the transcription factor and DllA. The necessary rran:-r'iptron factor is not blnding as well to the DNA, ..,..ding to reduced efficiency of transcription. The slde -:ain of aspartic acid can participate in hydrogen bond:r urth DNA bases, but the side chain of vahne (com: eteli-carbon-hydrogen bonds) cannot (see the figure :-.rri.). Tianscription factors do not bind to DNA ua .:ric interactions. An increase in hydrogen bonding r.:\reen the factor and DNA would lead to enhanced .:,..liscription (not reduced transcriprlon). An inabiliq. . -:he transcriprion lacror to bind ro DNA would lead ro . ,- transcription of the gene; since there is a 1ou. level ol : rtresslon, binding of this factor to the DNA must be ,:urring, although the affinity for the factor and DNA
:-:.s been reduced.

53

18. The answer is B. When region B is deleted front the promoter regron, there is an increase in or-erall
expression of the reporter gene (the second and lasr hnes of the figure indicare thrs). Removal of any other region leads to a decrease in expression, indicaring that posrtive-acting transcription factors bind to those regions of DNA.

r9

The answer is C. When regron C rs removed fiom the DNA, there is a tremendous reduction in the expres_

sion of the reporter gene, indicating that thrs region of the DNA binds important positi\.e1y-acting transcriprion lactors for the expression ol the reporter gene. The answer is A: Cytochrome synthesis. The ll,oman has a form of porphyria. an inborn error in the biosprthesls of heme. The buildup of heme inrermediates is rvhat generates the symptoms observed. Barbrturates are degraded through the cytochrome p45O senes of enzymes, lvhich are induced by the drug. The enz)-mes require heme, so heme sprthesis is also rnduced, and whenever heme sprthesis is induced, the toxic intermediates get accumulated. The end product of heme degradation is biiirubin, and rf excessive bilirubin accumuiates, jaundlce rs the result. Both dohchol and ubiquinone synthesis are from isoprenoids, and not
related io the heme pathwal

C:C I -r
t-N

I -CHs - c-c-\cH" |

ort

H,(

C- CH"-C_

?:o H
,1,-

,-:mson of valine (V) and asparagine (N) side chains

:

sithin

-.;kbone.

.:taa!::=::-.--+:a

Chapter 7

Molecular Medicine and Techniques

future of diagnostic medicine is in molecular medicine and the analYsis oJ Dl{A and Po'{A for
The these teclmiques.
sites also shown. The insert was obtained from a gene, and exon I ofthe gene is indicated by the black box.

mutations and susceptibility genes. This chapter will test your hnowledge of the application oJ

QUESTIONS
Select the single best answer.

bp

x
_o

patrent displays increasing muscle weakness and delayed muscle relaxation. He shows a slack .1aw and drooping eyelids. The physician has noted a loss of muscie mass in ihe cah.'es over the years. A molecular analysis by Southern blot using a probe agalnst the suspectecl disease gene shows the following. The moiecular

o o O
LO

i.uro. for the

dillerence between the normal and dls-

ease gene is which ol the following?

If this vector plus insert were to be cleaved to completion with enz)Tnes S and X simultaneousll', what size
bands would be observed?

(A) 1,600bp,2,600bp (B) a00bp, 1,500bP, 2,300bP

(C) 40Obp, r,0O0bp, 1,300bP, l,5O0bP (D) 800bp, 1,500bP, 1,900bP

(E) B00bp,900bP, 1,000bP, r,500bP

Which labeled DNA fragment would work best for
D = Person with disease
tA

Ce ne d upl icat ion

(B) Gene deletion (C) TripleL repeal exPan>ion (D) Increased SNPs in the disease state (E) Differences in restriction fragment length po1;tmorphisms (RFLPs) below is a vector plus insert, with
Shown Questions 2 and 3 are based on the following diagram'

Northern blot analysis? (A) The 3O0bp HX restrlction fragment (B) The f ,100bp HS restriction fragment (C) The 500bp HS restriction fragment (D) The 1,8O0bp SS restriction fragment (E) The 800bp XS restrictron fragment
You are studpng a family that exhibits a rare autosomal recessive disease. The disease is caused by a point muta-

tion whrch abolishes an EcoRl restriction site in

genome. To genotype the family, one can

the

restriction endonuclease

amplily a 1 8kb

54

Moiecular Medicine and Technique-'

region of this area of the genome using polymerase chain reactlon (PCR), restdct the PCR products with EcoR1, and then separate the restrlcted DNA on an agarose gel' (A) Southern blot
(B) Southwestern blot

5t

(c) MicroRNA analysis

(D) Microarray analysis
(E) ELISAS

The results of such an experiment are shown be1ow, using DNA obtained from each of the flve famlly members. Which family members have the di'sease?

1.8 kb

A patient was given an initial ELISA screening test for HIV which was positive. A second, more specific test was then undertaken, which consisted of a Western blot. The samples that were run through the polyacrylamlde ge1 for thls test consisted of whlch of the
following?

nd

12kb

0.6 kb
':,a4a:,.

(A) Patient sera (B) Patient plasma (C) HIV RNA (D) HIV proteins (E) Extracts of patient white blood cells A patient has been diagnosed with abetalipoproteinemia. Such disorder can be determined by which of the following techniques using samples obtalned from the
a

(A) A and

B E

(B) C and (C) D

)

blood? (A) Western blot

(D) A, B, and D

(E) C, D, and E
H

)p

(B) Northern blot (C) Southern blot (D) PCR analysis

A difficult step ln the engineering of recombinant insu1in was which of the followrng? (A) Expressing the entire insulin cDNA in bacteria (B) Expressing the entire insulin gene in bacteria

(E) Microarray

.;$j

l'reDNA sequence represented by rhe gel shownbelow' '="''"' whlch was generated using the Sanger dldeoxy sequencing technique, ls which of the foliowing?

(C) Findlng bacteria

preproproteln plestze

to properly

cleave the

.insulin

ATCG
A and B chains to
come together
ce11

(D) Allowing the

appropriatelY

(E) Flnding the appropriate eukaryotic

express the gene

i.n

which to

",*1 "-"'"'

1,6-month-o1d has had recurrent bouts of pneumonia'

ur-rd a chest

X-ray shows a greatly reduced th).'rnus. Gene

therapy would successfully treat this disorder if the gene is placed in which ce11 type or organ?

for

(A) Muscle (B) Liver (C) Lung (D) Bone marow (E) Kidney
-.?=

;omal mutan the

A patient has been diagnosed with a particular forrn of .rrr.ar. Appropriate treatment of this cancer, however, requires knowledge of which molecular markers are being expressed by the tumor as compared to normal
cells of the same tissue. This is most easiiy accomplished

(A) ACTAACGCTTA (B) ATTCGCAATCA

(C) ATCTATCGATC

r.8kb

by which of the following techniques?

(D) GCCCTTTAAAA (E) AAAATTTCCCG

'-]t''

,{J-,'

A --

variety of patlent has seen her physician for-a 'piarnts over the past ttuttut years These lnclude

fevers which

".*,*orth, h.,

bruising' and increased feelings of anxto provide a iety and depression A Jolecular technique
easy

stlffu'-td go' farigue' lornt pain and (see the figure below)' sores ln rash on the face

to*t

is u'-hich of the .oi-rn.rrn,io,-t of the suspected diagnosis followlng?

@
(A) B and C (B) A and D (C) A and C (D) B and D

(E) C and D
14 gene X' one wants Harzing cloned the cDNA for mouse a human cDNA to use this cDNA as a probe to screen the human homolog of this gene'

ilra|

and isolate which of ihe folurrt riair^,ior-t conditions should be library? fo*"g for the initial screening of the

(A) Western blot

(B) Southern blot (C) Northern blot (D) DNA sequencing (E) RFLP analYsis

(A) High temperalure, lor'l'salt (B) Lor,v temperature, 1ow salt (C) High temperature, hlgh salt (D) Low temPerature, high salt (E) Low temperature, no salt

t5 A researcher

12' '-''

run on ce1ls from norTwo-dimensional PAGE gels were tlssue which has been on ii-"

i"r*t ,"^';;.; ;t.pn,.d '.-pf'^"r,ri"

^q s-

Y He perhas isolared the cDNA for gene uslng ,f"r. fol1or'r'lng Northern blot experiment
i'o'.,-' di'fferent tissues A potential ior this fincling ls which of the following?

-ui tirr,r.,

u"d also

affected by a certain disease state' as compared best describes spot A in the disease state? to spot B in the notmal

'u*t Which of the following

.c

^a\
,.-6

Normal

Disease

0r
SIZE

fe

pH

pH

(A) Lower molecular weight' same isoelectric protein (B) Higher molecular weight' more acidic more basic protein icl uigfr.t molecular weight' irl Su*. molecular werght' more basjc protein

points

iuj ao*.,

moiecular weight' more acidic protein

(A) Aiternative sPlicing (B) mRNA instabilitY (C) RNA editing (D) Different AUG start codons in different (E) Differential polyadenylation
16

tissues

'

be amplified by ff-,. reglon of DNA shown below is to use is whrch The appropriate pair of primers to
PCR. of the following?

time in the hosTwo babies were born at the same up' but their nametags may have been mixed
frta1,

Molecular Medlcine and Techniques 57 DNA fingerprinting was done to determine the parentage of the children, using Southern blots of restricted DNA and a specrfic probe. The basis for seerng differences between indir,rduals using this technique is rvhich rl the following?
r,r,ou1d be expected

ampiifled DNA by enzyme X, a carrier of the dise;... to exhibit tvhrch size fragmenls. :.. determined by ethidtum bromide staining of an agage1?

rose

\r

Chromosomal delelions

Disease

B) RrLPs C) Chromosomal translocations

D) TripleL repear expansrons

XXX

E) DNA methylation
5',

JIJ

ltt I

l<-oer.o---+

-- mtcroarray experlment looking at genes expressed b1. :. cell line both before and after differentiation ol the --ne indicated 15 potential genes which \\,ere upregu-

{_ 1.5 kb _______+ <-1.Bkb_--------->

Region to be amplified

simple technique to enable the scienrrsr ro ,-:termine the temporal order of induced gene expres. .rn is which of the following? i) Southern blot lr Northern blot Jr Western blot
--..ted.

l)

(A) 1.5kb only (B) 0.6 and 0.9 kb (C) 0.6 and I.5kb (D) 0.6, 0.9, and 1.5kb (E) 0.9 and I 5kb
Consider the following map ola genomic region of DNA, showing restrictlon endonuclease sites for enz),rnes X and Y You har.e a probe to this region (as indicated on the figure). A certain disease maps to this region of DNA, and creates a new restrlctlon site, Z, which is cut with restrlction enzyme Z. As a diagnostic tool, a carrier would exhlbir which bands when DNA rhar has been completely restricted u,ith enzl,rne X is run through a ge1 and a Southern blot rs perlormed utilizing the indicated probe?

RFLP analysis

F'r Microarray analysis

-r genomic clone has been isolated and is going to be -.sed for chromosome walking to obtain more clones
-- the gene. The regions of the clone to be used as probes r chromosome walking are which of the following?

XX I z Y
-! l' :
-''
BandC

AandD
A and E D and E

J,,no

l*35kb+
5.0

I <f
= Probe

kb --------+

BandD
studies a dlsease caused by a single point muta-

lre

,-,.n rn which a restriction enzyme site is gained by the ::ltat1on. To determine if someone is a carrier of the :-sease, PCR primers were generated which allowed a

- S kb fragment to be amplified across borh X resrric,--.n sites in the normal gene. After treatment of the

(A) 5.0kb (B) 1.0 and 5.0kb (C) 1.5 and 5.0kb (D) 1.5 and 3.5kb (E) 0.5 and l.0kh

58 '.1

ChaPter I

be cut rnto enables the DNA lo restrlction srte thatthe disease' as rs A is a carrier of the

ANSWERS
The ansurer is

ii"'rt.."t ;;;;;;t'

(: Triplet

ol myoronic has the about b1 a tripler ';,*ptorrrf somal dominnnt'o''ltotJ ''ro'gn''

repeat expans'on presence

The patient rePeat exPansion' d)strophy an aulo-

f-'ut t*" normal '"divrdual individual B individual'' carrier of the disease' not a

copies of the

"l']l1:,.';:l';ff,' ffi
'ht d"

il'il;';;;:.:lf'n:il:';;ffi .;;...''u
o-uu' o.o.ur
ro two chromosomes has

'5

The answer is D:

Arrowin{n:

l::::J:l'l'^tl"tL:
e\

tosether

"rll:o:ii:::r.,",';::l;;,,^,i*,,

enrs ro

:1ilIl1,*":i,|o.'':i;t; fot
conlain tl-'i' t"pu'-"to'-t
ease Myotom't

expansion *

U"'*n' " of the DMI #'t;illranslared ltcl9" g. I., r.., y, T',;.Jf ;Ti' [, :l!if and is located o *,T,:;."T"H:,, gene by a gene duplication' dystrophy * ""tl^"tta chanse tn a *"''i;; *u"utut ll:.':,"^f n' ' Iength pol; deletion tn '" SNP parterns' ..l'i'tf "ntes 'resrriction rhe triplet may be created bv morphlsms ('l;;";g;;"

disparienr to har''e rhe repeat to a'CTG triplet

::':1,',".?':ll t""}"';^i'f "" o[ the C-peptide) the clea'age "l '""1;" i;*;va1 occur in bactenot lo form mature i'-"uii'l would insulin' the A and ttt"*U'"^"t ria. Thus, i'-' *ulti'lg punfied' ttplt^t"ry in bacterla' B chains u" ptod""j approprlate condiand then mi"ecl togethtt
tions to allow the

such e,enrs) rhus'

U'*ina" bo'-'d' ro'totrrr i'nsulin One cannot express chains to lorm *n*tt do not bacterla' as bacteria
the enrire insulin
r:

''-'att

between the

:;,,il-i^ l*j*:1il,."fiT:
:Xii

gtlt '"

:lt*,
o

-"
at

rePeat exPansion)'

\ll J".i1:

I JJfi ;;;." J, i,.',.,ii"

ap p r

ri

erv'

900bp' 1'000bp' The answer is E: 800bp' tr""''"p :ltlt figure' the next Starring **l tr-'Ji '* ^t the insert' gent"'i""fa f" atthe X srte in
restnclion erating ^ next site i'

1'500bp'

and ar high leve1s'

DNAl";t"r"lr

'ht the next.cut site Molrng ufotg tf-t" nNA

t'ooool,t'oo long' i" the inserL' whlh is 800bp site in ;';i?" ls the X

+ 300) The

The answer is D: Bone

adenosine marrou'' The child has

SC1D (severe

rleaminase

immunoderic*"" ;"';;"i1.

rhevec,or,'qJi;;;ill:::::;::"i:T:,.ti:J'"1',,T; n ra'l d 'I:i" : J,;'i:"::'il : T ;; :l 1 *'' "e s seup to the '''tttoo add

il' :ll:lling ;rd F ;;:v: i.t1*$ :i*:,:f 'Uh:'iJili: runcblood

ce1ls

to'ol o"iitt"t;,-"t

combined

il

*.1ffi,?"il:t"il};;;'""sine
tional ADA

Bv.incorporating
stem
ce11s

i" b"":';;;;"*-tt1lt"tr-tt

for the

800bp' "ooiol'
total,'1'200bP'
The answer
Slnce

u''-ta

'

:oor'p'

;;fi

;,

ADA'

."!,',

'

tl'"

the DNA

f";il';'.""bt

restriction fragment' C: The 500bp HS Northern blot' p'Jit'" ;;"; "*"tb3 usld in a from an exon' '; bt olyined

is

and lvill now "*ptt"-'i'i' tistrutt"g tlt'gtt-'" ,n other effects of dt"*yud";;;i'-t" of blood death
sues
cells

";, "' and the matt

*,*::,ffi jl::3 ;:H ;:'l.T'::i; to [he toxrc

be resis[ant
of tlie

will

'-'ot

uaa'i""'i;p;;i#

'

as mRNA

-'*'";;- ;;:':'l

:::T:"tTt:ffi:'h;i;.'r" nry* tl" ;oobP ili *"'tt'"" n^91:::r:lj*jT::\#:

which
other Parrs oI the insert in a Northern blot hybridlze ro mnNa

Tl:'ln$i|T"fi[is within

The answer
:

I JlJ:.

Mrcroarray is D: Microarray analysis'

t:f"::lj]i E'

[ il:;l$':;;;.'
;*i

i:: Ut:i ita" trt*]^'

;;*. "..' (abour rl00

explai'ned

"rr'

"','fi to an arra) or known

iI ilI,:lL

ff

,it''

Th" answer is B:

carry a u"tf-t chromosomes must disease' so 'o*ut "tt"-ittt' the person to have the since the mutated g""t f"t 'ffiu;t;;iu be seen on the gel only one restrrcdisease destrovs a
mutatton

and

is auloSince the disease

:Tj:,:;';';;*; is) i1.,,,, t" tn: expressron levels of

l^*ll.rermine the
south-

" I l.. I i ""'J:'B"JI :i

iltitiii: ai:;n: in srngle

srare l-ffiT.T: ,"ffo,i'ffi.ao:^" ""a ""'*al r1.n11ue,i; *oura rrave

er:n blors cannot 1ar results.

,ion'naollli'^""-"' "":i'il:;1il:' lilt;i :":il i}in?.'#;'l,"i *'v*' rt'"'' Hff *?i,T: :i:;# ;*"?'*n: r"' i'.a'i'lfi ;'"i;'? YolX11 ;:T; whtt
allele' sPonds to the mutant

;;';jt;;;to

probes to try to obtain ;l:r"#n ii-oo atrt""t r'ioi' * whrch protern'P}T

""t

smt-

'the

sot"}'*"'tt],r'

,^;*o'tt;tu"t"d'
MicroRNAs regulare

siluatlon' is not relevant to this

v", ^u*, ::r::ju.1j:"t:tl,J (and mtcroarruY g:1:'

**"'''t^"*tt'"on'

but will not tell

b^"d' t;;"';;l;

'k

normal

al1e1e'

which contalns

;^;".;;.t;;ni'llts'qt

I n through examLne one prote

i'ffi

;"jffi

Molecular N4edicine ancl Techniques

59

.:ubody binding and cannot gi,.e rapid

.tr00 different gene products.

data

on

oYer

,,1O,

il:,::cscope slide

I I

In the Sanger technrqr-re. the sequence ls read from the bottom of the ge1 Lo the ,.trp of the ge1 (5' to 3'), going from one size ol D\A ttr the next largest size. This is due to the way in which the frasments are generated, as rndlcated in the figure be1ou.
The ansurer is B: ATTCGCAATCA.

A. Terminates with ddATP f--

l-7" .r.ri,I l,iD\.{t/

.-\

r6l
t,

ri,
t.linrl
DNA l.

:"
i18

DNAtemplate

3'5'

T PriTer

- G- C- A - C- T - A -

T-

(Nucleotides: 5'-

-primer-10-'11-12-'13-14-15-16-17I
I
DNA po'ymerase + ooATP and other normal nlrcleotides

-5 - - 3 --

L--r l:l'i *" l\

6-,i

\:;

lnrr

uurn \

-.- :.

,'

,l

I ocrp, acrp, orrpt

Besuttant *
porvnucieotides

5'11:::TTr_
.4./fy!l\
"

lo

\--' \_-, --r

orrli

I lE?fr1 \ l. D\-{ ,

_. _. _. o#p
-. -. -. -!

-. -.-c

17

ddATp

/isntl\, :,1|Tr i) ' o:ri jl lf/lnral lt I '{-_7

l

7a"i''.

B. lf synthesis is terminated with:

lf synthesis is terminated with ddATP
'10

ddGTP ddCTP
t11

ddTTP

- -,: mlcroarray experiment. -- -: irr known genes (actin,

t
SiTe of nroalrcts

Note hor.r,, DNA sequences correcyclin Dl, etc.) are spotted in an :- ,r mrcroscope sllde. Labeled cDNA lrom ce11 samples u''ill -.:r-ed to the DNA on the slide, and the results analyzed by . :r to determine the 1eve1 of gene expressron at each spot

(in nucleotides)

--'l

/",;
t

Polyacrylamide gel electrophoresis

18 17 16 15 14 13 12
11

l/ tr/
10 8

i I I

ts

16

The answer is D: HIV

proteins. The HIV

tests are

looking

Size of DNA fragment (in nucleotides)

Sequence of newly synthesized strand read from bottom of gel

in the patient's sera againsl So, in the confirming Western b1oL, HIV "l\-proteins. ::-rreins are separated by size on a polyacrylamide ge1, le proteins transferred to fi1ter paper, and the filter paper :aated wlth the sera sample. 1f the sera contain antibodies .: HIV proteins, they wr11 bind to the proteins on the fllter :-.per, and the presence of antibodies on the fi.1ter paper is -::an detected using second antibodies. None of the other :rrS\\-ers are appropriate for this tlpe o[ test.
--r the presence of antibodies

5_ACGTGATA*3'

1O+17

11,, The answer is A: Western blot. The patlent has lupus,

r,vhich is an autoimmune disorder. To test for the presence of antrbodies ln the patient's sera, one can use a Western blot, and run typical antigens (such as splicesome proteins) on the gel, and then determine if the patients sera contain antibodies against the proteins. ELISAs will also work for this assay A11 of the other techniques mentioned require examlning nucleic aclds,

The answer is A: Western blot. Abetalipoproteinemia :sults lrom the absence of apolipoproteln B in chylomi::..ns and very low density lipoprotern (VLDL), caused

whrch would not help in determining

if

autoimmune

MTTP, the microsomal triglyceride one isolates plasma (remor.rng the -:LLs), and performs a Western blot lor the presence of -.:olipoprotern B, one would see greatly reduced levels -... compared to normal individuals. As apoB is made .--.her ln the lntestine (apoB'l8) or the liver (apoB100), ,.:oB mRNA would not be present in blood ceils, so a -...rthern blot would not be informative. Southern blot . DNA would only work if there was a speciflc probe .:i \{TTP which would distingursh a disease gene from .-. rormal gene. Microarray, which measures mRNA lev. .. rvould not show any difference between normal and i sease states, as the apoB protein is still made in the . er and intestine; however, it cannot be incorporated -' :o chylomicrons or VLDL due to the lack of functional :.ITTP activlty PCR analysls suffers from the same prob-:r as Southern blotting.

r.. a mutation in
-:;nsfer protein.

antibodies were being generated ln the patient.

lf

12:'

The answer is C: Higher molecular weight, more basi( pro-

tein. In two-dimensional

electrophoresis, proteins are

separated first by charge (they are run through a pH gradrent, and when they reach therr isoelectric point, they stop migrating), and then by size. As indicated in the figure to

the questlon, larger sizes migrate more slowly and would be closer to the "top" of the figure. Thus, spot A is of a larger molecular weight than spot B, as spot B migrates

larther

in the sizing

portion of the two-dimensional

electrophoresis. The horizontal positions are related to the isoelectric points ol the proteins, and spot A reaches

its isoelectnc point at a higher pH than does spot

B.

This means that protein B is more acidrc than protein A. Overall, then, the protein expressed by spot A is larger and more basic than the protein represented by spot B.

60

Chapter 7

The answer is A: B and

(.

For a PCR experiment,

:'i

one requires pnmers thar will allow DNA slmthesis to occur across the region of DNA to be amplified. Since DNA polpnerase always s)'nthesizes DNA in the 5' to 3' drrection, the 3' ends of the primers must face each other and bracket the region of DNA to be amplified. Primers A and D will only a1low DNA s1'nthesis away ln the opposite direction from the region oflnterest, due to their orientation on the DNA template strand. The prlncipal behind PCR reactions is shown in the figure below'
Reqion of DNA to be amplified

The ansurer is D: Low temperature' high salt. Since the probe i.s from mouse DNA, and the sample is human DNA, low stringency conditions need to be used to a11ow slight mismatches in hybridization to be tolerated. Thus,

1ow temperature reduces thermai motion, and a11ows for mismalches, and high salt will reduce the potential disruptlve lonic interactions that mrght result from mismatches and alterations in backbone stfl-1cture. The combrnation of low temperature and high salt will provide the highest probability of hybndizatlon between mouse and human DNA.

3'Strand2 5'Strandl
Cycte

,r,

.{+

ttre answer is A: Alternative splicing. Si.nce

Northern

1
I + r

Strand'1 str?nfz

3'5r

I I I I Y
I

Heat to separate strands

Coot and aOO

blot ls being run, RNA from the various t.issues is run through the ge1 to be separated based on size. Seeing
dlfferent sizes of hybridizing bands, depending on the tissue of origin, wlth the same probe, and from the same animal, indicates that alternative splicing of the mRNA has occurred between the tissues. mRNA instability would lead to reduced 1eve1s of signal and not differences in sizes. RNA editlng wouid not alter the overall size of the mRNA; it would ;ust alter the sequence of the mRNA. Start codons are withln the mRNA; utilizing different start codons would not alter the size of the mRNA. Differential polyadenylatlon is only correct if it is a part of differential splicing, which is the best
answer.

pnmers

.--I

Strandl
Strand2

3'-

I ndd nearstabte I DNA v Pol!/merase s',@-*------r

-t
Cycle

5--=--T@r5| 2 r r+

Heat and cool

| I
'l'
.@_----

twith Frimers and

DNA Polymerase Presentt
I

strandl 3

+-r

Strand2

---__w ]* 5'*
I I

+r

"':t6.,,

The ansuret is B: RFLPs. RFLPs can be detected wlth a straightforward Southern b1ot. Chromosomal trans-

@--------------M

w--r
--..-M --=--M @F@ ---t

locations are too rare in the populatlon to be used for general DNA f,ngerprinting. DNA methylation patterns are epigenetic and can be variable amongst indir'rduais, so they are not sufflciently rehable to be used for DNA fingerprinting. Deletions and triplet repeat expansions are too rare and varlable in the population to be used as a general screen for DNA identification purposes.

Strandl

3'-

e;;'t vI ff5:,;lififl.
@---------.--M @--=l

.i?.r
''

w@
M

Once indivldual genes The answet is B: llorthern have been identified as being either upregulated or downregulated after differentiation, the DNA corresponding to the gene can be used as a probe in Northern blots using mRNA obtalned at various times after differentiatlon i's induced. In thls manner, the ti.me course of lncrease, or decrease, ln mRNA expression can be determined None of the other techniques will a11ow this information to be easiiy obtained. Western blots require antibodies to the proteins produced from the genes identifi.ed, which most like1y are not available. A microarray analysis is too

blot.

rw-r --.---wer@ @I
@
@

----------------

Strand2

r--.-_@l 5'ffi cycles4 | . io 20 '* |

l=----r

Multipleheating and cooling cycles

r--=]
'106

complex to easily determine the temporal order of gene expression. RFLP analysis does not address the question of temporal gene exPression.

Amplified DNA present in about

copies

Molecular Medicine and Techniques

6l

Tle answer is C: A and E. Chromosome walking (the DNA at --.-. as taking the "ends" o[ a probe

19

The answer

is D: 0.6, 0.9, and 1.5kb. A uirr,l:

and 3' ends of a probe) and using those as new r3s ln another library screen. The clones identlfied - ..--. second screentng of the library will hybrldize to -. .::id'' of the already ldentified clone, and should : -.ri rhe DNA both in the 5' and 3' directions from -:, ,.-ready represented by the first clone, as indicated .--: figure be1ow.

-. :'

wouid have one normal gene, and one disease gan.. whlch contains the new restriction enzyme site. Aite:
amplifying this area of DNA by PCR, and then cuttinr

with the appropriate restriction enzyme, thrs rndividuai would show a 1.5kb band from the normal
gene, and both 0.6 and 0.9 kb bands from the disease
gene.

@@
S;=c 1: use

u" a probe
Extends cloned region at the 5' end

Step

2: ,se

as a probe

Extends cloned region at the 3' end

5.0kb. This would be a noninformative result. Enzyme X would not cut at site Z, so there would be no difference between the normal and disease gene when the DNA is cut with enzyrne X, so both genes would give rise to a 5 kb plece of DNA. In order to distinguish between the normal and the disease gene, one needs to cut wlth two enzymes, X and Z; using the probe indicated in a Southern blot, the disease gene would then show a prece of DNA that is 1.2 kb tn size, while the normal gene would show a piece of DNA that is 5 .0 kb in size.
The answer is A:

Chapter 8

Energy Metabclism? Overview

The use of energt is a key far biorhemics,l pathways to ptroceed. This chapter cavers the basic tenets of biocbemic{tl enirgy utilizatiott, storage, and production, as well ss tlze bodv,s
efiergy needs"

Given thrs same patients eating habits and hfesr1.1e, whrch of the following best describes her metabolic
state?

QUESTIONS
Select the single best answer.

(A) She is gaining weight (B) She is in calorlc balance (C) She is in the heahhl. range of BMi but is losing
weight

(D) She is in an unhealthy range of BMI and is losrng

weight

Questions I to 4 concern the following case. you see in your office a thin, anxious woman who is concerned about her weight. She is worried that she may have a parasite causing her to lose weight. She stands 5'5" tall (f .67m) and weighs r0r lb (4s.85 kg).

(E)

She has a rapeworm and needs 1ab tesring

An esrimate of her body mass index (BMI) is whlch of

the following?

Circn ihe lollou ing reacrion: A + B 5 C + D AG.' = +15.5kca7mol And [A] = 5mM, lBl = 4mM, [C] = 0.5mM, and [o] =

2.5mM under
1.98

ce11ular conditions, what

is the overall

(A) (B)

14 16 18

Gibbs free energy change for the reacrion at 25.C (R =

10-r kcaUmoU"K) (rn kcaUmol)?

(c)

(D) 20

(E) 22

2 The same
ca1 day

patient then describes to you a rypt_ eating, u,hich consists of 2509 of car_ bohydrates, 10g of fat, and 1009 of protein. She denies any ethanol intake. She also exercises about 2Wday. Her daily caloric rntake is about which of rhe

(A) +t3.Bo (B) -13.86 (C) +15.:0 (D) -15.50 (E) +17. t:

ol

Con>ider the iollou ing reaction seque"rce:

ASBAG"'=+0.50kcaVmo1
B

following?

(A) (B)

1,250 1,500 1,750

l" D AG"' = -12.15kcaUmol 5 E AG'r' = +21.15kcaUmol

C AG"' = *15.50 kcaVmol

(c)

Under standard conditions, whrch intermediate would

accumulate?

(D) 2,000

(E) 2,250

(A) A

(B)

For the same patient, her daily caloric needs can be estimared to be which of the followrng?

(C) C (D) D

(A) 1,250 (B) 1,500

(E)

(c)

1,750

(D) 2,ooo (E) 2,250

Questions 7 and 8 refer to an unusual bacterium that has been shown to have a five-component electron transfer chain. Table 8-l shows the E.'values for these five componenrs.

62

Energy Metabolism
:.-. E i1

Ovelleu' 63

1l
. _n1n Llt)

+0.55V
+0.12V

Ivan Applebocl is an over\veighr accountant. u'iih ,.. height of 5'9" (1 77m) and au.elghL ol245lb (i I 1 -1kr' As a sedentary individual, his daily caloric nee d apprrr\imates whlch of the lb11oning?

. at-l17 ur
DH2

+0.03v

: -:-- )
-

-0.22V

. trut? Lt

-0.41v
flor,v in thls bacterium is u.hich of

:ier of electron

(A) 2,;00. el (B) 3,000 ca1 (C) 3,500ca1 (D) 4,ooo ca1 (E) 4,500 cal

- --!r\\ ing? - --- :ransfers to B, u,hich transfers to C, lvhlch trans::s ro D, u,hlch translers to E = :ranslers to D, r,vhich transfers .:s to B, r,vhich translers to A - :iansfers to D, u.hlch translers ::-: io B. rvhich Lransfers to A :ransfers to E, whrch transfers '::s to D, which transfers to C : ,:ansfers to C, which transfers ::s to D. which translers to B
to C, r,vhlch transto E, r,vhich transto B, rvhich transto A, rvhich trans-

12

Which of the follor,nng diet pians u.r1l allou, Mr Applebod to lose approximatell. 5lb/month assuming he cloes nor
increase hrs acLir.it
v-?

(A) 2,000cal/day (B) 2,450 caVday (C) 2.900 caVday (D) 3,350 caVdat' (E) 3,800 caVday
Questions 13 and 14 refer to the following figure indicating an energy curve for an enz)ryne-catalyzed reaction:

-:

' --

bacterial strain relerenced in the prer.ious quesamount of energy available from transporting .l .lectrons across this chain is u,hich of rhe follor,v. = 1.98 x t0 I kcaUmol4( and F= 23kcaVmol-V)
:r.-

I kcal/mol
O)

- -l kcaVmol

--

kcal/mol - - kcaUmol
- --.r kcaVmol

o
ut
C

-' --r1rrr\. - \\-hich

::

male medical student is 5'9" tall and welghs of the follolving diets will aliow mainte-. -.i the current r,veight and also fa11s u.rthin currenL --.rritdl guidelines?

Progress of the reaction ------>

.-,rout l00g fat, 50g of ethanol, 100g of prorein, -,rC 300 g ol carbohydrate
:-rLrut 80 g fat, ).25 g of proteln, and 310 g ol carbo.

drate

13

Which letter represents the Grbbs lree energlactivation?

of

--,rol1t 125g fat, 85g protein, and 350g of carbo.

drate
14

r.rout 80 g fat, 60 g ethanol, 1 00 g proteln, and 3 t0 g ::.:bohydrate .rrrut 50 g fat, 25 g protein, and 225 g carbohydrate -a

Which letter best represents the difference in

between the substrates and products?

energl-

t::r of calories? -r1)g protein, 100g fat, 100g carbohydrate, 25g

Lrne

ol the follow'ing diets provides lor the largest

15

Consider the reaction sholr,-n belolv

If

lA] =

>.00mN,1,

. -ianO1

:-rq protein, 1009 [at, 1509 carbohydrate,25g

r,vhat u-ou1d the concentration of D have to be to allow. thrs to be a favorable reaction under these conditions? A + B 5 C + D AG"' = +8.65 kcaUrnol

lBl = 2.50mM, and [C] = 1.25mN1.

-:

. -:]anO1

protein, 125 gfat.50g carbohydrate, no ethanoi rr.i g protein , 75 g fat, 125 g carbohydrate, 20 g g

. -:anO1

:ig

protein, 50g fat, 125g carbohydrate, no ethanol

(A) <0.125pM (B) <0.43 pM (C) <4.3nM (D) <4:nv (E) <5.0ttM

;E tei

h.< i:roea.sed from about 1.5 miles/day to l0 miles/day.

I *tt*.s rco" 5h erplains that she had made the cross_country rrm ar h"t high school, and over the past 3 weeks her running

li' a6g-ilisnist as she is concerned about los_ mrts TBgLr- ]ht is i 7" raII and q,,slghs l28lb, dovrn from l35lb
gnmc Ero

rrufil,lrmli

r mri t - ndtr to rhe following

case.

A l5_year_old

,8

Calculatton of the basal metabolic rate (BMR) for mor_ obese individuals using standard methodology is ofren incorrecr due ro which of the following? (A) Underestimation of calories consumed (B) Overestimation of calories consumed (C) Preponderance of metabolically active adipocytes

bidly

tG

(A) 8oo (B) 1,ooo

Assuming that the parients weight loss over the past 3 weeks was equally distrlbured over rhat time perlod, how many calories per day was she deficient in her diet?

(D) Preponderance of inert adipocytes (E) Reduced metabolic need of the muscles

t9

The BMI rs most 1ike1y to 1,re1d incorrect data for which of the follomng?

(c)
(D) (E)

1,200 1,400 1,600

(A) An anorexic 25-year-oldwoman (B) An obese 50-year-o1d man

(C) A normal appearing 30-year-old man (D) A 30-year-o1d female bodybuilder (E) A slightly overweight 42-year-o\d, biochemistry
professor

17

After learning rhat her diet was deficient in calories, the runner decided ro make up the deficit by eating equal amounts (ln term of calories) of carbohdyrates and pro_ teins, but no fat or a1coho1. How many grams of carbs and proteins would she have to add to her cliet in order to stop iosing weight? (A) 100 g of each (B) 150g ofeach (C) 200g ofeach (D) 100g of carbohydrares, 200g of protein (E) t00g of protein, 200g of carbohydrate

20

Assume that beer contains 5olo wt./vol. ethanol. How

(about l7 ounces) of beer contain? Choose the closest answer to your calculated value.

many calories denved from alcohol wouid 500mL

(A) ioo (B) 140

(c)

180

(D) 220

(E) zoo

Energy Metabolism Or-en

.r.'. 6;

-r\

9145

"
:

ihe answer is B: 15. The BMI is calculated as the weight .:-. person (in kg) divided by the height squared

(in meters). Thus, for thrs patient, the Bltl1 is c-... . 45.85 diuded by (f .67)'], which is i6.44 The B\11 -. -,.:- *. for body mass index, and can be used to estlmai. '- -fat content. A value of <18.5 is considered undenr. -:.'..
values between 18.5 and 24.9 are considered ur ::,. normal range, vaiues of 25 through 29.9 are consicie::: overwerght, and rralues of 30 or greater are consi.ic:lobese. Values of 40 or more are considered n.rorhLdr'.. obese. whereas values between 35 and 40 are consrcered clinically obese. The formulas in order to perfonl: these calculatlons are summarized in a figure abore. The second figure allows one to utrlize a graph to cr1culate the BMI.

- :. on of body mass index (BMl)

Weight

(kg) Height (m)2

Kilograms = pounds + 2.2 Meters= inches + 39.4

:rnan who is5'4" talland weighs 134|b

a BMI of 23.5.
'134 lb + 2.2 =

61kg
(1

The answer is B:

1,500.

Carbohydrates conrain,l caVg.

64 in. + 39.4 = 1.6 m;

.6)2 = 2.6

protein also contains 4caUg, and fat contains 9ca1,/g (because it is more reduced than either protern or
carbohydrates). Given the patients dret, she is consuming (250 x 4) + (10 x 9) + (100 x 4), or 1,490caUday The answer is C:

BMI-

61 ko

"

2.6 m

=23.5

1,750. The

BMR can be esrimared

How
Herght'

Lo calculate the BMI

BMI (body mass index)

by multiplying the \\.eight (in kg) times 24 callkg. r,vhich is assuming an energy use ol 1 callh/kg. lvlulti3A

18.5

25

plying 45.85 times 24 yields 1,100cal/day Her meta-

bolic need can be estimated by multiplying her BMR times an acitivty factor (hou. acti\.e the indivicluaL is). For someone who exercises ZVday (moderately' aciive) the activrty factor is 1.6, so her daily caloric
needs are 1,100 x 1.6, or 1,760 cal/day. More accurate representatlons of the BMR can be obtained from using the formuias in Table B-2, although for the purposes of this textbook the approximation of 24 cal/ kgiday wr1l be utihzed for both males and females ol

so 75 loo''u . - .,.'-:Lght in pounds.

-a:
e

al1 ages. J.l,o"oo"l*u,io,?*":",**;t;"r:"'

.::m

used to calculate BMi, knowlng the height rn inches,

8-2. Equation for predicting BMR from body weight (W) in kg

Males Females BMR kcal/day 60.9W Age Range (years) 0-3 3-10
10-18
1

{e

Range (years)

BMR kcal/day

54

610W-51
22.5\N + 499
12.2\N +746
14.7Vtl + 496

22.1\l'1 + 495

175W + 651 15.3W + 679

1'1.6W + 879

8-30

30-60
>60

8.7W + 829
10.5W + 596

13.5W + 487

-::

: -lealth

Energy and protein requirements: Report of a Joint FAOAfuHO/UNU Expert Consultation.Technical report Organization. 1987.71. See also Schofield et al. Hum Nutr Clin Nutr. 1985;39 (suppl).

seriei no. f24. Geneve

66

Chapter

fhe answer is D: She is in an unhealthy range of BMI and is Iosing weight. The BMI of 16.4 places the ind,rvidual in an underweight situation, and she is currently consumrng fewer caiories (1,'+90) per day than
she requires (1,760), which will lead to weight ioss, and further exacerbate her underweight condition. You have a diagnosis, there is no reason to pursue further testing' She can be counseled on hor.v to gain weight.

The answer is A: +'l

Recali, LG = LGo'+ RTln this reaction, AG = 15.5 + (1 98 x ([C]tDl/tAl[B]), so for 19 r)(298) ln (i 25120). Thus, AG = 15.5 + (0.59) 1n (0.0625). AG = 15.5 - I.6+ = l3.86kcaVmo1 As AG is a positive number, under these conditions, the reaction
rs

3.86.

24 cdldaykg). Being sedentary, the activity factor is I 3, for a total daily caloric need of 2,480caVday. Current nutritional guidelines indicate that no more than 30o/o ol one's daiiy calones should be fat, so the maximum caloric intake for fat should be 750cal, rvhich is about 80 g of fat (fat contains 9 caUg). These data alone eliminatl answers A and C. Answer E has insufficlent total calories ('i50 from fat, 100 lrom protein, and 900 from carbohydrates, for a total of 1,450) for the needs ol the srudent, and can also be eliminated. Answer D contains too much ethanol (420cal out of a total of 2,780) and too many calories. Thus, answer B is correct, in which the 80 g of lat provide 720 ca|, the 125 g of pro-

tein pror.ides 500cal, and the 310g of carbohydrates

provides 1,2'|0ca1, lor a total of 2,460caVday
The answer is D: 150g protein, T5g fat, l25g carbohy' drate,20g ethanol, To anslver this question, one needs to recall that fat conlains 9 cd/g, ethanol 7 caUg' and protein and carbohydrates 4caVg each. Using these numbers, diet A contains 1,875 cal, diet B contains 1,875 cal diet C contains 1.625ca1, diet D contains 1,915cal, anc

still an unhvorable reaction.

D. The conversion of A to B is slightly unfavorable, but as soon as B is produced it will be converted to C due to the highly favorable B to C conversion (a hrgh negative AG). The conr''ersion of C to D is also highly favorable, whi.ch rvil1 lead to accumulation of D. The conversion of D to E, however, is highly unfavorable (hrgh positive AG), such that D rvill accumulate
The answer is D:

diet r1

ctrntains l,550eal.

under standard conditions.

The answer is B: E transfers to D, which transfers to C, which transfers to B, which transfers to A. The order of electron flow goes from the lowest standard redox potential (E/EH,, with a r''alue of -0.'+7V) to the i-righ.tL redox poientral (A/AH,, with a value of +0 5 5 V) ' Redox pairs wlth low redox potentlals are good reducing agents (they like to glve up their electrons), whereas

The answer is C:

3,500cal. Mr Applebods BMR can bt approximated as24 x 111.'1, or 2,673caUday. Since h< is sedentary, his actrvrty factor is 1.3, and has a dail'
caloric need of 3,475caUdaY.

t2

redox pairs with hlgh redox potentials are good oxldizing agents (they love to accept electrons). Thus, the ord..tt electron transfer would be E to D to C to B and then to A as the terminal electron acceplor.

The answer is C: 2,900ca1/day. To lose one pound c' r,veight, a reduction in intake ol approximately 3,500 cr' is required; thus, a loss of 51b will require a reduction c ' 17,500 cal over the next 30 days (one month). Dir''iding 17,500 by 30 yields 583 caVday Since his normal intak' (to maintain weight) ts 3 ,47 5 callday, 3 ,47 5 - 583 yielc' 2,B92caUday.

t3
The answer is C: 47kcal/mol. In order to answer thls queslion, one needs to use the Nernst equation, which equates overall changes ln redox potential to Gibbs free energl The Nernst equalion is AG"' = -nFAE"', where

n is ihe number ol electrons transferred, F is Faraday's constant, and AEo'is the change in redox potential ln this case, AE"'ts equal to 1.02V (the diflerence between trav-0.47 and +0.55), and n = 2 for a pair of electrons thus becomes eling through rhe chain. The equatlon
LGo'

The answer is B. The energy required to increase lh' energy siate of the starting material (indicated by A t: the diagram) ls the Gibbs free energy of Activation Th' change in energy states of reactants and products is ind" cated by E, while D shows the maximal energy chans' lrom the energy of actlvation to the energy 1eve1 of t1:: products.

14

The answer is

=-(2X23X1.02) = -'17 kcaVmol

The answer is B: About 80g

fat, 125g of protein, and In order to answ-er this ques3l0g of carbohydrate.

energy staie of the starting material (indicated b;' ' in the diagram) is the Gibbs free energy of activatio:' The change in energy states of reactants and produc'' is indicated by E, while D shows the maximal ener: change from the energy of activation to the energy 1er ' of the products.
The answer is C: Recall, AG = AG"'+ RTthe reaction to be favorab ' In order for

E.

The energy required to increase ih:

tion, one first needs to calculate the basic daily energy needs of the student. At 1751b (79.55kg) one can estlmate his BMR as I,910caUdav (79.55kg multiplied by

15

<4.3nM.

(tcltDl/tAl[B]).

Energy Metabohsm Overr-re'".

67

, -

,.:: :le negative. If one solves for the concentratlon

.:

- \l[B]) Therefore, 8.65 = -(1.98 x 10jX298) l--Dl/[12.5]). This reduces to -14.66 = ln (D/10), , 'oLi) = [D] in mM. lDl = 4.3 x 10-6 mM, or
l9
is

-.rred for AG = 0, then any concentration iower . rne calculated will be sufficient to al1ow for a . -\G. Thus, when AG is set to zero. AGo'= -RT In

inactrve, and rf an rndividual has a lot of adipose i-..,.. the contrlbutron of the adipose tissue to the overalj :1.-:. will lead to an o\/erestimate of the energ,v needs ..- - .

indlvidual. It is not related to the amount of cal!ri-.. consumed, nor does rt relate to use of the muscles. sir,:.
the BMR is estimated for energy use during resr.
The answer is D: A 3O-year-old female

bodybuilder. Tre

-he answer is C: 1,200. Each pound of weight

,'.-:nt to about 3,500cal. The runner had lost 71b, -,.r1 deficit of 24,500ca1 As she lost that weight - iay's, her daily deficit, rf evenly distributed over . -.:se ol the three weeks, was l,l67caVday

ffi -le
Ca

llr:rilir'

answer

is B: 1509 of each. Both proteins and .-..irates contain 4caUg, so to make up approxi-

i:

'' 'Crates would have to be 300y'day Since the ' :r '.iefltS to spirt the calories equally belween pro. : Carbohydrates, l50g ofeach is a better answer ' -.:-Ltting the 300g into l00g of one nutrient, and , . :enothernutrient.

BMI is an estimaie of the "fitness" ol an indir,rdual. and is calculated by taking the w-eight, in krlograms, ano dividing by the square of the heighL, in meters. \hlues between 18.5 and )4.9 are considered the normal range. while values above 24 9 fal1 into the preobese and obese categories. Body builders, r.vhether male or female, har e an increased muscle mass for their herght, which adds welght. Thus, body builders u,il1 have an lnflated B\11 r,vhich rs not indlcative of their fat content (it reflects therr muscle mass instead). The other indir.rduals listed r,r.ell mostly fit the criteria for a valid BMI determinarion
20

The answer is C:
500

(59100mL) Alcohol contains 7caUg, so 7

180. With alcohol at 5ok wt./r-ol.. mL of beer would contain 25 g of alcohol

25 =

-lt

l75cal. Beer also contains some carbohydrates, so its

answer is D: Preponderance of inert

adipocytes.

A11

-,.:s of the BMR utihze the werght of the individ.--',\.ever, adrpose tissue is prlmarily metabohcally

total calonc content u,ould be even higher than the t 7i due to the ethanol alone. In contrasl, 12 ounces ofa cola product t1prca1ly contains 150 ca1 (none from ethanol).

Chapter 9

ffi#trffiG#Ha#ffi

ffiffi#

%reHBffiEreffi

fuE#tr EemeeroHEBffi
it:-rludirrg str:ixttl #ltsst?"zfiti's, :*j-sLtii'!i, glr-,"t*g*;=-. pht: sylwtid-.ifireosii* J, *td steri:id tu*r;c; *:2.* sigt:*-iirz'g" Btcttust tfuevt is aru*th*r *tr,yttt' {}tz {*t{'!'{. sfErtr.ri tt' *-*s tlur;ri*t. p ra{}*v rzy s sp rcipts.ii j' t',: l*-ii-sz1 t * {{:tr,{ {{ src disttt-ssrd ;* t;z*t th,*pttt: Thi"s riz*l:'Lr j i#1 rI:tltt ilfl r s gentr *1 * sll {: {t s t>! sigzi:2l- t|' 4i .x-i'|'rt{: ! i {}i'i..
Tfuis riz-ttpte i- #J\,{i"s I,izt 1:*sit= ,si sig'tzr.;i {r*-::'*ititiii:t'",
\\ratery diarrhea, vomiting, and leg cramps. The mole., 1ar basis ol his disorder is rvhlch of the follou.ing?
"

(A) Inhibition ol protein synthesis (B) Inhibltlon ol a stimulatory G protein

(C) Actlr.ation of a stimulatory G protein (D) Inhibition of an inhibitory G proteln

(E) Activatron of an rnhibrtor,v

G protein

QUHSTg{}i-;$i
Select the single best answer.

A 40-year-o1d man, with a BMI oi 37, has a lasting bl.glucose leve1 of 165 mg/dl-, a value that has been sleac increasing over the past 5 years. His HbAlc 1evel is E

A middle-aged female has developed darkened, smooih,

velr,ety skin patches on the neck and in her axrllary folds (see the f,gure below). The molecular basis for this disease can be which of the fol1or'r,ing?

Analysls of his blood, using Western blot technolt: demonstrates the presence ol antibodies agalnst t- . lnsulin receptor. One consequence of these antibod ., is which of the following? (A) Doll,rLregulation of the insuiin receptors (B) Upregulation of the insulin receptors (C) Enhanced release olpancreatic glucagon (D) Enhanced glucose uptake into muscle ce11s (E) Actir.ation ol adenylate cyclase Considering the patient in the previous question, \\,hic of the following medical disorders would you expe, this patrent to also have? (A) Lor,v blood pressure (B) Polycystic ovary sl,ndrome (PCOS)

(C)
Note the darkened skin along the neck.

Fatty iiver-nonalcoholic hepatrtis (NASHnonalcoholic steatohepatitis)

(D) Hypourlcemia

(A) (B) (C) (D) (E)

(E) High HDL cholesterol

Dyslunctional glucagon receptors Dysfunctional EGF receptors Persistently stimulated rnsuhn receptors
Persistently inhibited insulin receptors D1 sfunctronal thyroid hormone receptor
Consi.denng the patrent in questlon 3, which of the follou ing body tlpes would you expect this patient to have?
-

(A) Body builder

A 4-year-o1d boy went to the shore with his family and ate clams for dinner, but shorti,v thereafter der.eloped severe

(B) Central obesity (C) Obesity mostly centered in the hips and breasts (D) Ta1l and slender (E) Ideal body weight

68

Hormones and Srgnairng N'lechanrsms

-.r - \\ith a good appetlte has been losing weighi - ,. ieveloped lesions and blisters on his [eeL and -, --,r analysis demonstrates hyperglycemia under - .- and fasting conditions. A possible explanation -::. s\ rnptoms is which of the followlng? -. . :cagon-secreting tumor of the pancreas - -nsulin-secreting tumor ol the pancreas - ss of glucose-6-phosphatase activity

69

ll

A researcher r'vas im.estigating the mechanism of autrl:. : steroid hormone receplors. He created a chimenc rei.-:-

tor that contained ihe estrogen ligand-brnding domain and also ihe iestosterone DNA-btndrng domain. The transaclivaiion domain was also from the testosterone receptor. When this chimeric recepror was expressed in
eukaryotic cells lacking estrogen receptors, which ol the following u,or-rld you expect to occur when estrogen is added to the celis? (A) Increase in tyrosine kinase actit ity (B) Activation of PI{A (C) lnduction of estrogen-specific genes

- :.::tive glucagon receptors :.:ruclion of the G cells of the pancreas

' srgnal transductlon u,,as mutated such that its -- ---:rain was no longer functional. Alter adding " :. :.r this cell 1ine, proteln kinase B couid no longer
- -.:ited. The protein which u,as mutated is r,vhich

:e

has been generated in which a proteln impor-

(D) Induction ol testosterone-specific genes (E) Inhibltion of estrogen-specific genes

,.,1orr,.ing?

l2 A 17-year-o1d girl was seen by her family

a-

::-F

-l'-kinase
.-.ts2.

physician dr-re to a lack of menses. On physical exam the girl appeared to be r,ve11-der.e1oped, but u.ith a sinktng lack of pubic hair. Blood u,ork Lndicated very high leve1s of testosterone, u,ith eler.ated estradiol ier,els as u,ell. A karyotype of the patient is shown belolv The underlying biochemical delect in this patlent ls rvhich of the following?

'::.-i:cher was stud),1ng a tumor in n'rice that

u,as

, , -.; bv transfection of a rrector, which led to overpro, , : of c-ras within the tumor. The researcher wanted - ' -rdd a second expression vector that would coun-. , -:r. eilects of the overexpressed c-ras. Which one -: -r11or,vlng genes should be investrgated?

*e

&&:asi 1 r, il* a,lia

{f#

t-P
_:

ls
St4 d

:---::r-old boy had frequent infections and a number of . ..::onia e pisodes. BIood work showed greatly reduced . ,. -.i both T and B l1.mphocytes. X-rays showed nor-

-...i

&*:t
. .:','mus der.elopment. The primary mutation in this .::t rs likely to be which of the following?
.lr,rtated JAK

Afird.i6adt

*ti

{yr *& Y*
10 11

sq

s!9**:; 13

t&.s, ,sS 8&;*

iai l,t

rS ., tif,S! 16

+'e 11

:i
18

protelns

14

15

'rrated STAT proteins lelective cytokine receptor component

l.lutated S\trAD proteins
l,.lutated ubiquitin ligase

&* l9

.*6

&

St *i

*&i
21

..
22

*; t

t ;

20

::id-class sprinter, while in the starting blocks wait: ---r ihe race to start, stimulates which of the follow: :ruscle proteins ln response [o hormone release?
., tr-rosine kinase
Go,s protein .- Goi protein -- Gq proteln :-{\1P phosphodiesterase

The karyotype

liom the patient

(A) Inability to slmthesize estrogen (B) Lack of estrogen receptor (C) Lack of cortlsol receptor (D) Lack of androgen receptor (E) Inability to s)nthesize aldosterone

E-?;?=:

70

Chapter 9

l3 A variant cell line was

discovered that would not respond to any form of TGF-B, yet both type 1 and type il receptors were present and functional on the cel1 surface, as determined by ln vitro experiments. A potential
lead to thls phenotype?

mutation in whlch one of the following proteins can

(A) (B) (C) (D) (E)

Constitutive activation of the EGF pathr,vay in bone lnactivation of the TGF-B pathway in bone Constltutive activation of the FGF pathway in bone lnactivatron of the FGF pathway in bone Constitutrve activation of theJAIVSTAf pathway in
bone

(A) SMAD2 (B) SMAD3 (C) SN,L{D4

(D) JAK1 (E) TYKI

A 52-year-oid man suddeniy collapses and is rushed to

the hospital, where it is found that his blood troponin-C
levels are elevated. An angiogram indicates an 87olo blockage of three major arteries io the heart. If these arteries

t4

62-year-old man visits his physician

for

feeling

fatigued. It is noticed that the spleen is enlarged and slightly tender when palpated. He has abdomrnal pain and ech).,rnoses on his skln. His blood pressure rs high. Blood work indicates an abnormally hlgh red blood ce11, white blood cell, and piatelet counts. A bone marrow sample from the patient is shown below. The suggested treatment is phlebotomy to reduce the thickness of the blood. This dlsorder can result from which of the following?

were examlned at a molecular 1eve1, excessive smooth muscle cel1 proliferation would be noticed. This prollferation has occurred over the past 25 years due to the secretion and action ol which of the following factors? (A) VLDL

(B) HDL (C) FGF (D) PDGF (E) EGF t7 A mouse fibroblast ceIl line is treated with an siRNA targeted toward MEK. When such celis are treated with
a growth factor which works through a tyrosine kinase receptor, which one of the following effects would occur?

(A) Activation of ERK (B) Activatlon of myc

(C) Activation of phospholipase (PLC-y)

(D) Activation of los (E) Activation of STAT

t8 Tieatment of fibroblasts with TGF-B and TGF-u will

al1ow such ce1ls to grow in soft agar, a property of trans-

formed cel1s. This occurs due to whlch of the following? (A) Activation of ce1l growth (B) Activation of oxidative phosphorylation (C) Activation of collagen slmthesis

(A) A loss of function of JAK2 (B) A gain of function of JAK2

(C) A loss of function of SMAD'I (D) A gain of function of SMAD4

(D) Inhibirion ol ras expression (E) Inhibition of LETS slrLthesis

(E) Constitutive t5

t9 An example of an autocrine stimulatory pathway

which of the following?

is

He12 act ivrty

A 3-month-old child of normal parents presents with shortened arms and iegs, a large head with a prominent forehead, a curved iower spine, and bowed lower Iegs. The child is in the fifth percentile for height. The child's conditlon is most likeiy the result of whrch of the following?

vessel growth within a tumor (B) Glucagon release from the pancreas (C) Insulin siimulation of glucose transport into adipocytes

(A) Biood

(D) Injured smooth muscle

injuries

ce1ls

producing PDGF
vessel

(E) Platelets secreting PDGF at internal blood

7l
- .. - -' :- . :.. * --. ll :I^-:. -1...'*.r .-1.1 . ir ;:,.i. i: .li i - . s. :. -.:t:.->--\-T-: eliher rhe serlne or threo_ - * - r . -,- ;r rh.,:ph.,n l..rred pho.phor; lation -: .
: a . r- I l. ..r- f

- .- j -.r Ii:'-.. --'fl ,rr :icli\e prolein. ln cerla,n .. .r . .: - .,r J. ti\.lti-Tg n utJtion is .ound in this -.-:,:::ti.:s:he protein acti\-e in Lhe absence of any

(A) Ator (B) StoA (C) Ttoe

(D)

rn.spno:\ larion er ent. \\ hrch one oi the iollourng aminr acid mutarions is most likelv an acti\-ating muiationl

Ator

(E) StoY

'f
72
Chapter 9
causes cholera (Vibrio cholerae) produces a toxln tha: ADP-ribosylates a stimuiatory G protein, leading tc

ANSWHffi.S
The answer is D: Persistently inhibited insulin receptors. The patlent has acanthosis nigricans, whlch results lrom insulin resistance. One way in u'hlch insulin resistance

permanent acttvarion

can be obtained is rhrough a persisrenr inhibirion of insulin receptor actiyiry Alterations in the glucagon,
EGII or thyroid hormone receptors r,vi1l not lead ro rnsu_
1in resistance.

GTpase acrlviry). This leads rc constant cAMP production, u,hich in rurn leads to the secretion of sodium and water info the lumen of the intestine, producingtvatery diarrhea. Ilnot treated, deh\dration rapidly resuks, and dearh can occur. The figuie belorv outlines the aclion ol cholera toxin in inrestlnai
epiLhelial ceiis.

tron

ol the intrinsic

of the G protein (r,ra inhibt_

protein. The bol.has contracted choiera. The bacterium r,vhrch

G

The answer is C: Activation

of a stimutatory

t$ater contaBinatsd

$ith

srroreffa

Vibrios c*lonize small rniestine

I
Brndrng of Chotera toxln
I

Choiera tox!n

?
il
tf il

i&irasellular

E[$rrHTlx$fi ?3? il tl
Ittilt
tl

*h*l*ra texin
{A subuniti
ADP riba'sylaticr *f 6-Br0t8in

t_

&

J$$

*
lnhibition ot STFase astivity 0f G-p.stein

D,,:*"Lt-Ltt
Orr-l*GD\

l"',

Fsrsjsts*t activttion 6t ad*nyiyl *yclas* by GTP

_{LJ LJ _rft___, .-t.=-i

:

*
Ma$sive socretion

*f Ha' and l{n0

ffiT.P -\l -1*-fNa , H,o Cholera

:ir

lA,ffil

*l[-cr*trl
t\.

(A

toxin subunit)

)
SEYERE OiABRHEA

.*.*.--".*--*/

;j

*
Pf;HYO*ATION

sH0cr(
*
T}EATH The bacteria contatning the cholera toxin are ingested and travel through the dlgestive s),stem, and r,vithin the intestlne their toxin exerts rnd elecrro\-ie ioss. Dehydration ani hypor.olemic shock can result.

effects. u'hich leads to seyere wrLr'r

1ts

-fre a:srer is A: Downregulation of the insulin receptors.

.

-- ..- ---.- ;. ..--: -'.t-..<--.:-.:''-.nL'- : - ' --= -: -:J :aaa::.ri. rancinq trr dour-Lregulalton r :-:r -r Ci-.. ..r: :nus kepl in a persistentll'1o$' - -..:-::,,-: s::.:.. l;unling their abilit)'to be stimu- .:.s',- :r The Lack oi Lnsuhn srgnaling leads to . .--:-::: iiucose ierels. These alteraitons are not - : -i:r-_ii: in qlucagon secrellon or acti\-alion olade- . . -"--,...e ,iince muscle cells have dour-Lregulated : - - .::3piors. the muscle ce11s are not stimuiated
--:-- -':

.-.-.:..,

- ^ .-- . --_...--

:-j:-.-,:..-

:--j.-..:--

: -

=lucose lrom the circulation. The antibodies ---ri arlori- lor upregulatlon of insulin receptors.

The answer is A: A glucagon'secreting tumor of the pancreas. The patreni has a giucagonoma in rii.]icr the pancreas is constantll' secretlng glucagon, leading to consiant gluconeogenesis and fatty acid oxidatlon. despite the presence of adequate glucose 1eve1s in the blood. This leads to w-eight loss and hyperglycemra, as the liver ls pumping out glucose despite normal blood glucose levels. An insulin-secreting tumor would lead to hypoglycemia (stimulation of glucose uptake from the blood even when blood glucose 1eve1s are 1ow). As g1ucagon is released from the cr-cells of the pancreas, loss of cr-cells lvould not allou' for glucagon secretion, and
these symptoms rvould not appear. A lack of liver gluca-

-ire answer is C: Fatty liver-nonalcoholic hepatitis (I'IASHrccal<ohol ic steatohepatitis) . The fuil insulin resistance - -:-:--. consisls ol diabetes mellitus 2, central obeslty,

gon receptors would not a11ow-hyperglycemia to occur (hypoglycemia w-ould be obsen ed instead), as would a lack of glucose-6-phosphatase. The lesions on the leet
and legs are due to nonenzymatic glycosylation of nerves

' :r :mslor. lou- HDL, high tnglycerides, high PAI-I

-'-:

marker of inflammar- il uric acid, and NASH. A11 of these lead to total - - :,a.rosc1erosis. PCOS is part of this constellatron : r-::oms. but occurs only in women. These insulin-,. -i,-,:--, patients most hkely also har.e sleep apnea, but -. .. ::obabiy lrom the obesity and not from the insu- r.j s:ance. The causes of NASH (fatty liver, ieading to
gen acti\-ator
a

:' :.

inhibitor- l,

and small blood vessels in the periphery leading to reduced circulation and feeling in the extremitles. The answer is D: Pl-3'-kinase. Protein kinase B is recruited to the membrane, along u'tth the phosphoinositide-dependant kinase, through binding to PIP. in
the membrane. The PIP. ls generated from PIP, through the actions of Pl-3'-kinase. If Pl-3'-kinase had lost its SH2 domains, this protein would no longer be able to bind to phosphorylated IRS-1, and in turn be activated by the lnsulin receptor tyrosine kinase activity Proteins containing SH2 domains are capable of binding to other

-.-.:.-s' are multifactorial, but lnsuiln resistance ls one

,

- -:

,har can lead to the development of NASH

Tbe answer is B: Central

obesity. The patient

has insu-

l::-SlnFIl.

.. ::slstance sy.ndrome, which is a polygenetrc disor-. :sua1ly brought on by gaining werght. Classically, :-',- -n resistance s1T-rdrome has a central obesiq- pat.-. The patients BMI is 37 (clinically obese), so he -.-. d not be tal1 and slender and is not in the ideal - :-. u-erght range. A body builder could have a BMI of - rut this would be due to muscle mass and not due , . ';ipose tissue, so he would most 1ike1y not be insuhn

proteins which contarn phosphotyrosine residues. in the absence ol Pl-3'-kinase activity, proteln krnase B could not be activated (see the figure below), as both
PKB and PDKl (phosphoinositide-dependant kinase 1) require phosphatidylinositol trisphosphate in the membrane as a docking station, through the proteins pleck-

strln homoiogy domain. None of the other

choices

listed (ras, raf, GAB GRB2) require P1-3'-kinase activity in order to be activated.

Pl-3,4,5{risP
Activated
PI 3-kinase

PH

domains Phosphorylation and activation of PKB by PDK 1

Dissociation

PDKl

PKB

Answer

f=

74

Chapter 9 The answer is A: GAP. Overexpresslon of ras leads to transformation because there is insufficlent GAP (GTPase-activatlng protein) to inactivate the ras in the ce1l. This can be overcome by adding GAP to the celis, such that the ras to GAP ratio is cioser to one. GAP can thus inhj.bit ras activity and block transformalion. This

will not work with oncogenic ras, as oncogenic ras has obtained a mutation whlch has inacttvated its bullt-tn
GTPase activity. Thus, GAP cannot actiYate oncogenic ras and cannot block ras function. SOS is a guanine nucleotide exchange protein that leads to ras activation; it cannot turn off actlvated ras. Raf, fos, and PI(A. do not play a role in regulating ras activily. A summary of these activlties is show'n in the figure above. The answer is C: Defective cytokine receptor component.

{Y\ GDP GTP ";\ t Pi

\ r''

t (-.|;.1 lcrPlcf'
/

\(errase //,'' \/l \ r-------r,/

/ r---r--__l '-ri

r?l Iuo,
I GrP ]l -*T-

Effector response

Many cytokine receptors contain multiple subunits, and some subunits are shared by multiple receptors. The boy has X-hnked severe combtned immunodeficiencl s;,rrdrome (SCID), which is a defect in a cytokine receptor component, the y-chaln, common to a large number of cytokine receptors. Lack of this component renders most cytokine receptors inoperable, leading to lack of response to cytokines and inabllity to mount delenses agalnst an immune attack. The defect ls not in the TGF-B/SMAD pathway, nor in the JAK/STAf family of proteins. It rs also not related to a ubiquitin ligase, which targets proteins for proteolysis by the proteasome. 10

\rTt foo, IcrP]l

lli , _ rr!I

*
Effector response
Panel A indlcates the regulation of the nonnal ras proiein (p2 1). Panel B indicates the lack of regulatlon of the oncogenic form of the ras proteln, whose GTPase actir,ity has been ablated, such that GAP bindlng to p21 cannot reduce the actir-ity 1evel of the mutated ras protetn.

The answer is B: A Gcrs protein. The sprinter has released epinephrine in anticlpation of the race. Epinephrine brnds to a heptahellcal receptor, which is linked to a stimulatory G protein. Upon epinephrine bindlng the G protein is activated, and the Gus subunit binds GTP and travels to, and activates, adenylate cyclase, which raises rntracellular cAMP 1evels. Epinephrine binding to such muscle receptors does not result in an lncrease of tyrosine kinase, inhlbitoqG protein, phospholipid-specific G protein (G.), and cAMP phosphodiesterase activlties. A summary'of different types of heterotrimeric G protein subunits is given in Table 9-1.

Table 9-1. Subunits of heterotrimeric G proteins

Gcr

subunit

Action
Stimulates adenyl cyclase

Some Physiologic Uses

Glucagon and epinephrine to regulate metabolic enzymes, regulatory polypeptide hormones to control steroid hormone and thyroid hormone synthesis, and some neurotransmitters (e.9., dopamine) to control ion channels Epinephrine, many neurotransmitters including acetylcholine, dopamine, and serotonin
Has a role in the transducin pathway, which mediates detection of Irght in the eye
Co

cr.; Gcr(s)'

cr,"; Gcr(i/o) (signal also flows

Inhibits adenylyl cyclase

through
o,;G*(t)

By subunits)

Stimulates cGMP
phosphod iesterase
)

o0,,,; Go (g/1'1

Activates phospholipase

Epinephrine, acetylcholine, histamine, thyroid-stimulating hormone (TSH), interleukin 8, somatostatin, angiotensin Thromboxane 42, lysophosphatidic acid

d,r,.,.,G0 (12113)

Alters cytoskeletal elements

,There is a growing tendency to designate the heterotrimeric G protein subunits without using subscripts so that they are actually visible to the naked eye.

Hormones and Signallng Mechanisms 75

-\e

:-

- .:r bound to the receptor, the receptor would , : nd to testosterone response elements in promot.i1. genome (the DNA-binding domain was spe-- : the testosterone receptor). The sterord hormone

answer is D: lnduction of testosterone-specific genes. - :rnle ric receptor was created such that when

receptors do not have lyrosine kinase activity, nor do they actl\.ate PICA.. Srnce the cells berng used for this experiment lack the estrogen receptor, there would not be any ellect of adding estrogen on estrogen-specific gene expression. Tl-Le chimeric receptor is shown in the figure belour

Domains of the steroid hormone receptor

Dimerization sites

lnhibitor binding sites

NLS t, /7
C

to

Transactivation domain (TAD)

DNA binding domain (DBD)

Ligand binding domain (LBD)

Testosterone domains

Estrogen

ln the presence of estrogen, the receptor will be activated, and will bind to the testosterone binding domains in DNA, and will also interact with transactivation proteins specific for the testosterone receptor.

fhe answer is D: Lark of androgen receptor. The patlent . . .-:lrbitrng complete androgen insensitlvity syndrome -.-S'. an inability to respond to androgens due to a lack ,:'.-: androgen receptor. Thus, whiie genotlplcally a -',, .. and producing normal levels of male hormones, the

- *. .annot respond to the hormones, and female sexual . -::j..cie nstics develop. Androgen rynthesis remains nor-, . hou.ever,

-cr-eLs

whlch leads to high circulating testosterin the patient.

transcription thctors. Once phosphorylated, the specific SMAD factor dimerizes with a common SMAD, SMAD4, Ior transport to the nucleus and blnding to the appropriate response elements in DNA. A lack of SMAD4 would account for the total lack of response to TGF-B, as all signaling through TGF-B requires this component. JAK and TYK work with cytokine receptors, not wlth TGF-B receptors, and SMADI and -2 are specific receptor SNL{DS; loss of either of them would diminish certain types of responses to TGF-B, but not
all responses to TGF-B. The pathway ol TGF-B signaling is shown below.

The answer

is C: SMAD4. The TGF-B receptors

to initiate phosphorylation of
SMAD

r^

together

rGF-B

_________>

1.TGF-B binds

2. Type ll receptor

to type ll receptor

phosphorylates type I receptor

3. Activated type I receptor phosphorylates R-Smad

4. R-Smad complexes with Co-Smad and

migrates to nucleus

t4

The patreni has poiycythemia veta, a myeloprohferative disorder, whlch has recently been shown to result from conslitutive JAKZ actir.ity. Under normal conditrons, JAK2 is only actir,ated when an appropriate cytokine binds to lts receptor. Once actlvated, JAK2 phosphorylates STAI
The answer is B: A gain of function of

lAK2.

protelns, which translocate to the nucleus and initiate gene transcription, leading to a proliferative response.

prohlerate a1'ar in ine absenre .': a sii:r..-. Sl.l-'l ::-' terns transmtt sLgnals from TGF-B srgnairng "ni ..:: :-, involved in this dlsorder. Her2 is the EGF r...D:whlch also does not play a role in this disorder F more about this case see Kralor.tcs R et al \ Enr Med 352'.177 9-2005). JAK-STAT signaling rs outlin:in the figure below

-:

::..,

--:

-: :

---r-

: -r.,r -,-.: .r-.

-,:.-

-:-:

-:

\5

2. JAKs phosphorylate each other and the

receptor phosphorylates STATs

t5

fhe answer is C: Constitutive activation of the FGF pathway in bone. The patrent has achondroplasia, the
most common form of short limb dwarfism in humans. This disorder is due to constitutive activation of FGFR3 (a receptor for FGF), which particularly plays havoc with the cartilaginous growth plate in 1imbs. Fu1l details of how constitutive activation of this pathway leads to all of the phenotlpic effects of this form of dwarfism are not yet fully appreclated. This disorder ls not related to alterations rn the EGII TGF-p, oTJAVSTAT pathways; rt is unique to constitutive activation of the FGF pathway vra FGF receptor 3. (For more on this case see Horton, WA, Ha1l, JG and Hecht, JT, Lancet 2007;370:162)

cell line. Effects which are upstream to MEK activation

will stll1 occur: effects downstream of MEK actiyation will be biocked. MEK actlvates ERK, which leads to alterations in gene transcription, such as myc and fos.
Tyrosine kinase receptors do not activate STAT proteins (that occurs with cytoklne receptors). PLC-y activation occurs upon receptor tyrosine kinase activation, and bindlng of the phospholipase to the receptor via its SH2

domains. Activation ol PLC-y does not require MEK aclivation. The ras-raf pathway leading to the activatlon of myc and fos is shown below.

Active Raf

1 1

Ras

t5 The answer is D: PDGF. During the deveiopment of

an atherosclerotic plaque, PDGF is released by platelets at the site of injury, which aids in the stimulation of smooth muscle ce1l proliferation as the damage is being repaired. PDGF is also synthesized by smooth

MEK

MAP

kinase@
Transcription

in an autocrine fashion, to further stimulate their own proliferation. FGF and EGF do not appear to play a role in the development of such plaques. HDL is a cholesterol carrier that removes cholesterol from tissues, and VLDL is the lipid carrier released by the liver for delivery of triglyceride to the tissues. (See Raines EW Cytokine Growth Factor Rev
muscle cells,
2004;15:237.)

/\
AP-1\g (iun and fos) \\ \/ 18
faclor@
tnaultion
ot myc and fos

Cell proliferation

,7 The answer is (:

Activation of phospholipase (PLC'y). The silencing RNA will ablate MEK expression in this

The answer is C: Activation of cottagen synthesis. The comblnatlon of TGF-cx and TGF-B leads to collagen production in f,broblasts. The collagen is secreted into

Hormones and Signaling Mechanrs:::-. 7 7

the cells

...: solt agar and provides an adherence polnt for .,:cts

the

\L\D prcnd are n.- ' recepto: rrder. (Fc:

-::oblasts, which ailorvs them to proliferate. The celis . , not grow if this adhcrence poinr is not present. The
of TGF-ct and TGF-B do include activation of cell .:.-)\\-th, but Lhis can only occur in solt agar if collagen -,.. been s1,r-rthesized and secreted. The factors do not - -'ck ras expressron or rnhrbit LETS (fibronectin) syn-

\
.s

Engl

outline:

muscle cells both secrete and respond to PDGF ---.. tumors secrete FGf; they stimulate endothelial cr : - liferation, which aids rn angiogenesis (the tumLr: ,:.--: are not responding to the FGF-the endothelia. :.--. are). Glucagon release from the pancreas is an endoc:::. process (the target for glucagon are cells far fr..n ::-. pancreas), as is insulin stimulation of glucose trans:.:.

','.

:srs. Oxidative phosphorylation rs also not directly

.,

-:cted by these growth factors.

in muscle. Platelets secreting PDGF ls not autocnn.. .1,: the platelets do not respond to PDGE The dilfere:::.
between endocrine, paracrine, and autocrine stimli tlons ls shown in the figure above.
--.-

i:
)l

The answer is D: lnjured smooth muscle cells producing PDGF. Autocrine stimulation is defined as one cell both ::,:.ting and responding to a grow'th factor. During the -.'.-aLopment of an atheroscierotic plaque, the smooth

rm

A. Endocrine
Hormone secreted into blood

. activatic:. : activatlr':-

Blood vessel

:h leads
r

i:

c and fos \T protelr-.

T"rS"t*lb@
B. Paracrine
Secretory cell

/ actlvatic:. -ation, an:

\ia its SHqurre ME:. e activaiio:

The answer is A: A to E. The conversion ol an r-.'.nine residue to a glutamate residue results in the inse:tion of a negative charge into this region of the pr..tein (the same overall effect that phosphorylation has The introductron of the negative charge would 1e;.; to slmilar conformatlonai changes [hat occur u'he:: a phosphate is introduced in this part of the prorer:r Thus, the glutamate side chain will mimic ihe eifect. of phosphorylation. Conr-erting a senne to an alanin: does not lnsert a negative charge into this area ol th. protein (the side chain of alanine is a methyl group thus, the required conformatron changes cannot occur The same rationale applies for converslon of a threonine to an aianine: the alanine side chain cannot b. modified to conLain negative charges. Conversion ..the alanine to a threonine introduces a hydroxyl group to ihe region, but the hydroxyl group has a high pK and lvill not deprotonate at physiological pH. Thrs is simrlar to the serine to tyrosrre conversion: t) ros na also has a hydroxyl group, but the pK" is too high tc Iead to significant deprotonation and introduction ol .1 negative charge into this area.

lnterstitial fluid

Adjacent
target cell

C, Autocrine

Target sites on same cell

hesis. Th. to collage:

ecreted int-

Receptor

" Hormone or other signal messenger

Chapter

1O

Glycolysis and Gluconeogenesis

This chapter quiTTes the student on the metabolic aspects af glycolysis and the synthesis of glucose, gluconeo gene sis. Yarious asp ects of diseases relate d to carbolqdrate metabolism are clso reviewed in this chapter
below An enzyme that may be defective in thts child
is which one of the following?

QUESTIONS
Select the single best answer.

Anaerobiosis leads to lactate formation in muscle due to whlch one of the following? (A) Inhibiting hexok.inase by giucose-6-phosphate (B) Provrding 2,3-bisphosphoglycerate for the phosphoglyce romut ase react ion (C) lnhibiting pyruvate kinase by pyruvate

(D) Providing substrate for glyceraldehyde-3-phosphate

dehydrogenase

(E) Inhibiting phosphofructokinase-1 by AMP

-Lthesis In muscle, under anaerobic conditions, the net syT of AfP starting from one mole of glucose derived from muscle glycogen is wh:ich one of the following?

(A) Fructose-1,6-bisphosphatase (B) Galactose-l-phosphate uridyiyltransferase (C) Galactokinase (D) Glycogen slmthase (E) Fructokinase

(A) 1 mole of AfP (B) 2 moles of AfP

(C) 3 moles of ATP (D) 4 moles of AfP

(E)

5 moles of ArP

A 2-week-old newborn was brought to the pediatrician due to frequent vomlting, lethargy, and diarrhea. Family history reveaied that the child never seemed to eat weil, and had only been breast-fed. Physical examination revealed an enlarged liver and jaundice. The pediatrician was suspicious of an inborn error of metabolism and referred the child to an ophthalmologist for a slitlamp exam, the result of whlch is shown

A7-year-old gir1, who lives on a farm, started to have shaking and sweating episodes. Upon physical examlnatlon, she was found to be hlpoglycemlc under fasting conditions (fasting blood glucose was 50 mg/dl) and posrtive for ketones in her blood and urine. Her growth curve is normal. Further analyses showed no other metabolic abnormalities. Probing further into her history, in the absence of her parents, revealed that one of her chores was to collect eggs from the chicken coop every morning, and she had gotten i.nto the habit of eating one or two raw eggs every mormng. This had been golng on for the past 6 weeks or so. A reasonable explanation for her laboratory results is which one of the following?

78

--

-,:

::--

:---::

.r::

\-\DH\,\D
genesrs

-.

--

!.

i(..

: r.li

_.ilI.

ratrc,

rC) The hrgh NAD-AiADH ratio impaired gluc.',nrc(D) The high NADFIIiAD* ratro impaired gluconeogenesis

--:i ::..dlLalion ---.::-i -er els trf electron acceptors in her system, -- :-r .. le d,,rced glucose production
.

(E) The hrgh NAD*,rNADH ratlo impaired glycolysis

A 3-month-old girl is brought to the pediatrician due

to lussiness and lethargy According to the parents, the baby was just fine until the mother needed to return to r,l,ork, and the baby r.vas being switched lrom breast milk to bab,v foods, formula, and lruit luices. At that trme, the child cried while feeding, sometimes vomlted, and had been lethargrc. The babys appetite seemed to have u,,orsened. The parents thought that if only formr-r1a was used, the baby u'as better, but they really could not remember. Whrch possible enzl,me defect might lead to this case presentation?

bout ol five days of severe nau. . :rr ,ng. lori'-grade ler.er, and diarrhea. This con,..i ,riflicted a number of people in Mr Smlths -: --,-:er recor-erlng Irom this drsorder, Mr SmiLh . '-. r-,i he cor-rld no longer drink mrlk before going '-.. -..s he became flatulent, his stomach hurt, and , : -.-.- cier-e1op diarrhea. 1l he did not drink milk, ..: -.-lditions did not occur. He had never expcri:* -rcSe problems belore the affliction. A possible ,,r-.,..irrrn lor N,Ir Smith's problem is u.hich one of the

' : :-

-.--.nrh' had

'.

-rql

.lrlhr.rnical disruption of the intestinal epithelial -.-1.. leadrng to reduced transcription of the galac-

(A) Galactokirrase (B) Fructokinase

(C) Aldolase

lilnase Elene l,l:chanical drsruption of the intestinal eprthelial ::11s. leading to reduced transcription ol the fruc,-r1lnase gene

(D) Hexoklnase

(E)

Glr-rcokinase

l.l-:c1'ranical disruption of the intestinal eplthelial

A thin, anxious woman, who is 5' 6" tall, welghs 92 1b. Blood u.ork rndicates a glucose level of 70 my'dl under

into the cel1s I'lechanical disruption of the rntestinal epithelial -r1ls. leadlng to loss of the glucoamylase complex rom their surface \lechanical drsruption of the intestrnal epithelial :111s, leading to loss of lactase from their surface
::11s. leading to loss oflactose transport

fasting conditions. Her liver is using which of the fbllou'ing as precursors Ibr glucose productlon under
these conditions?

...medjcs bring a patient to the emergency depart.--.:t because he was found unconscic'rus in an alley Dassers by. The man was unshaven and dishev..--:d. and appeared to be about 40 years old. Blood ., :..ho1 levels were found to be 0.25ok and blood :.-.rose levels 32 mg/dl. IV glucose was initrated, .:j Lhis enabled the man to regaln conscrousness, :..nough he was stiil inebriated. While conscious, ., eistorl, rer.ealed that the man \\'as a chronic alco-

.,,

(A) (B) (C) (D) (E)

G1ycero1, lactate, and leucine

Fatty acids, aianine, and glutamine Glycerol, lactate, and glutamine Glycerol, fatty acids, and glutamrne
Lactate, heme, and lysine

drinklng for the past 2 weeks, with nothing ,- eat. Analysis of liver enzyme levels in hls blood ::','ea1ed normal readings. Assuming that his liver is .-i11 lunctioning normal1y, why rs this patient hypo-.i cemic?

rlt

--iic. and as far as he could remember, he had been

A 50-year-old man has been trying to lose welght, but he enjoyed eating so much that he lound it difficult to do so. He then reads about a product in the popular press, which guarantees that he can lose weight, as caloric intake due to starch ingestion will be reduced (a starch blocker). The over-the-counter product that he buys is clalmed to lnhibit which of the following enz).r'nes?

(A) Pancreatic trlpslnogen (B) Pancreatic lipase (C) Salivary amylase (D) Gastnc amylase (E) Intestinal enteropeptidase

80 .lO

Chapter 10

hypertension, does nor have dyslipidemia, and does nor have antibodies directed against islet cells. This alteration in glucose homeostasis may be due to a mutatlon in which of the following en4rmes? (A) Pancreatic glucokinase

A 2S-year-o1d male deveiops diabetes, as nored by con_ stant, mildly elevated hyperglycemia. His father had similar s),Tnptoms at fhe same age as did his paternal grandmother. This patient is not obese, does not have

l2

The sprthesis of one moie of glucose from two moles of lacrare requrres six moles of Afp Which one olthe follow_ ing steps requires Afp in the gluconeogenic pathway? (A) Pyruvate kinase (B) Triosephosphare isomerase

(C) Glucose-6-phospharase (D) Fructose- 1,6-bisphosphatase

(E) Phosphoglycerate kinase

(B) Pancreatic hexokinase

(C) Liver glucokinase (D) Muscle hexokinase

l3 An important producr of the oxidation of the

enz)rmeS?

(E) Inresrlnal glucokinase

major energy source to provide energy for gluconeogen_ esis regulares which of the followlng key gluconeogenrc

bodvls

l.lr

A 3-month-oid girl with developing cararacts is shown to contain a reducing sugar in n.."r.irr., but the glu_
cose oxidase test was negative. She has had no problems eatlng, and her growth curve is at the 60th percentile. Fasting blood glucose tests show normai levels of circu_

lating glucose. the following?

A likely enzrye deficiency is which of

(A) PEPCK (B) Fructose-1,6-bisphosphatase (C) Glucose-6-phosphatase (D) Pyruvate carboxylase (E) Pyruvate kinase

(A) Fructokinase (B) Hexokinase (C) Galactokinase (D) Galactose-1-phosphate uridylyltransferase (E) Aldolase

l4 An individual with a BMI of 34 was

physlcian to eat less and exercise more. The prti.r_rt took this advice to an extreme, and has not eaten for 48 h. Which of the following best describes the patient,s activity and phosphorylation srare of the following key liver enzymes?

advised bv the

Phosphof ructokinase_

Glucokinase
Low Low
Low
No No
Yes

Activity

Phosphorylated?
Yes

Activity Phosphorylated? Activity

Low
Low Low Low Low

pyruvate Kinase phosphorylated?

Yes

(A) Low (B) Low (C) Low (D) Low (E) Low

No No
Yes

No
Yes

Low
Low

No No

No
Yes

No

15

Which of the following changes in

occur within of insulin?

enz)..rne acri\,1ty

h of a type 1 diabetic taking an in;ection

will

Liver PFK-2

Liver

PFK-I

Muscle Glucose

Uptake

(A) Active

kinase

Activekinase lncreased Activekinase lncreased Activekinase Increased

Decreased

(B) lnactive kinase

(C) Active phosphatase

(D) lnactive phosphatase Activekinase Decreased (E) Active kinase lnactive kinase

Glycolysis and Gluconeogenesis

8l

F '

-.'-'--,1-i t11utans,

found in dental plaque,

produces

19

Your 20-year-old male patient received a medLc;., discharge from the US Army. He has had muirrple
episodes of lightheadedness, sweating, fatrgue, tremor. and intense hunger. He had one seizure. During tri o

-:

:.atment in the dental office can slou'the pro-- acids. resulting in the accumulation of which

-:l

: .
-

-- -. -:se-6-phosphate

of these episodes, his blood glucose w-as 40 mg/dl Which ol the followlng tests could help you diagnose
his problem?

--:-.se-1,6-bisphosphate
-.

. ' ::raldehyde-3-phosphate -:
I.-.lhoenolpyruvate
:,,--1!
-.spho glycerate

(A) (B) (C) (D)

Fastlng blood glucose

HbAlc

a meal containing carbohydrates, the mono.,: ies must be absorbed from the intestlnal lumen. -:.msport is dependent on r.l,hich of the following

Noncontrast CT scan of the abdomen Blood glucose and insulin leveis measured while he was symptomatic (E) Determlning the presence of islet cell antibodies

l,lH-

,r-rriporr.,

Under condrtions of hypoglycemia, the liver is not utilizing glucose as an energy source due to which of the
following?

.,r.-.:ose-6-phosphate dehydrogenase

-.:'okinase
- :---..nde transporter .:. . K* ATPase

iA\ A lou K., for glucokinase (B) A high K, for glucokinase

(C) An rnhibited, phosphorylated PFK-1 (D) An activated, phosphorylated PFK-1 (E) A reduction of glucose transporters in the membrane

',

..1

muscle PFK-2 is not regulated by phospho-

- ::. but heart muscle PFK-2 is. In the heart, - .::rlrr)'lation of PFK-2 leads to what effect? , -::ranced production of fructose-2,6-bisphosphate ..:luced production of fructose-2,6-bisphosphate

l:gradation of fructose- 1 ,6-bisphosphate

:-lreased turnover of PFK-2 :lreased transcription of PFK-2

82 I

Chapter 10
regenerating NAD+ for use in glycolysis, specificallr_ ;.. a subsrrare lor the glyceraldehl,de_i_phosphate det_,.,

ANSWERS
The.answer is D: providing substrate for glyceraldehyde3-phosphate dehydrogenase. Uncle. anaerobic concli_ trons, the NADH generated b1, the glyceraldehyde_3_ phosphate dehydrogenase siep accumulates. Norma11y, the NADH would transfer rts electrons to mirochondrial NAD*, and the electrons r,vould be clonated ro rhe elec_ tron transfer chain. Hor,vever, in the absence of oxl,gen
Glucose

increase 2,3-bisphosphoglycerate 1eve1s, nor does it alie_ viate the lack ol NADr under these conditictns. pyruvatr

require 2,3-bisphosphoglycerate, anaerobiosis does n...

clitions. Similarlli w.hlle phosphoglyceromuLase doe.

does noL explain lactale formation under anaerobic con_

drogenase reacrion. While hexokinase is mhibrrej t, its product glucose-6-phosphate, rhis allosteric eftc..

\ \\

NADH+/

\*-rATP

\ Py"vate tr/ \ ,/ NADH<\ Lactate Acetyl CoA Anaerobic + TCA glYcolYsis

cycle is generated by the glyceraldehyde S_phosphate ,NADH dehydrogenase reaction; in the absence ot oryg"n, NnO. regenerated by the lactate dehydrogenase iJuciion. rn .is the presence of oxygen, NRDH Oonatei its etecirons to tne electron transfer chain (regenerating NAD+), which eventually donates the electrons to oxygen, forming water.

activator of phosphofructokinase_1; however. this actr_ vation does not relate to iactate formation under anaero_ bic,condltions. The figure summarizes these key pornrs outlined aboYe.
2

inhlbired b;. pyruvate (ATp and alanine are rhe allosteric inhibitors of this enzyme). AMp rs ar:

kinase is not

The answer is C: Three moles of ATp When glycogen produces glucose via the acrion of g11,cogen phosphori_ iase, glucose-1-phosphate L prod.,c"d.

the electron transfer chain is not fllnctioning. Thus, in the cytoplasm, the ler.els ol NAD* decrease to the point that there would be insuf_ ficlent NAD- available ro allow the glyceralclehycle_3_
as NADH accumulates

inhrbiting glycolysis. To prevenr glycolytic inhibrtion,

lactate dehydrogenase r,l.ill converi pyrllvate ro lactate.
Glycogen
I

phosphate dehydrogenase reaction to proceed, therebr,

verted to two molecules of py.rp1.n1., four moles of ATp are generated and one is utiiized at the pFK_l step for the net production of three moles of ATp Tr,vo moles of NADH are also produced, bur those are urilized by 1ac_ tate dehl.drogenase to reduce pyru\rate to lactate (anaer_ obic condrtlons) such that NAD* can be regenerated lor the gll,ceraldehyde-3-phosphate dehyd.Jg..ur. r,.p. A smal1 amounr of free giucose r,vrll te rjeased from glycogen by the debranchlng enzyme (about 5olo o1 rhe total); lor thar glucose, the net yieid is two moles ol ATp (since hexokinase has to phosphorylate the free glucose to glucose-6-phosphare), but since the majoriry of glu_ cose released is in the form ol glucose_i_phosphate,
three moles of ATp is the better
are outlined beiow

A, ,hi, i,

.or_r_

u.i-.r.

These reactrons

2-phosphoglycerate

+----+

phosphoenolpyruvate

I
Glucose 1-phosphate

t
I

+ 3-phosphoglycerate
,ArP
'1

,ADp -J

,Arp,l
pyruvate

+
Glucose 6-phosphate

*l

2NADH.

r, al
Lactate

Fructose 6-phosphate ATP

ADP

t I

. ,3 bisphosphoglycerate
2NADH

^:^':1
*

NAD'4

,n"*t_,
4 t 3-phosphate

2NAD_

\{

'

Glyceraldehyde

4l

t
I I

The flow of carbons is from glycogen to lactate under analroOit conditions.

Fructose 1,6-bisphosphate

+
Dihydroxyacetone phosphate

Answer 2

L-

Glycolysis and Gluconeogenesis answer is B: Galactose-1-phosphate uridylyltrans' ':'ase. The child has classic galactosemia, a defect -.-.,i.tose-1-phosphate urid,vlyltranslerase. Due to ,, -:r:mulation of galactose- I -phosphate , galac-,.e is rnhibited, and free galactose accumulates

83

-te

need of oxaloacetate lor giuconeogenesis, acetyl-CoA derived from latty acid oxtdation increases, Ieading to ketone bod), formation. Avidin does not affect NAD* or FAD ler,'els, nor does it interlere rvith coenzyme A or
vrtamin C. The answer is E: Mechanical disruption of the intesti' nal epithelial cells, leading to loss of lactase from their surface. Mechanical disrupiron ol the intestlnal eprthelial cells (as brought about b1' acute viral gastroenteriiis) leads to a ioss ol cel1 surlace enzl'mes, but iaclase rs the most severely alfected, as Lt rs present al the lor'vest leve1s on these cells. While glucoam,vlase is also iost, its actir.ity is in vast excess ofu,hat is required and rts partial loss does not allect rts ability Lo hydrol1.ze gLucose-glucose linkages (it does not hydrolyze lactose). A lack ol lactase means that the lactose in lhe diet passes undigested through the sn.rall intesline to the large inlestine rvhere the bacterial flora melabohze the lactose, producrng gases and acids that drsrupt the osmotic balance betrveen the lumen ol the bor,vei and the cel1s lining lt. This leads to \\'ater secrelion by the ce11s into the lumen of the bor've1, resultrng in dtarrhea. Lactose is not directi)' Lransported by intestinal epithelial cel1s (its components, glucose and galactose, are, afler hydrol,vsis ol B-l,a hnkage betrveen the trvo sugars), and a mechanical di.sruption of intestinal cells does not alter transcription of gaiactokinase and fructokinase.

'

ihe blood and trssues. The accumulatron of in the lens ol the eye provides substrate ---.e reductase, converting gaiactose to tis alco.:r (galaclitol). The accumulation of galactitol
-->c-

: .r ;1n osmotic imbalance across the lens, lead- . -.iiaract lormation. Additionalll', lhe increased . ..--1-phosphate, at very high ler''els in the
r,'

rrcks phosphoglucotnutase actlYlt)', result-

.:letfectir.'e glucose prodr-rction from glvcogen :l-rr\-iase degradation of glycogen rvill procluce .-i-phosphate, but this cannot be converted to

,-o-phosphate

if pl'rosphoglucomulase acLivrnLbited). A clefect in galactokrnase vu'i1l iead

Lrf

with cataract lormatron, the feeding problems assocraled r'vtth cias,-. rclosemla (associated with the accumula[ion -:ose-1-phosphate) are observed in nonclassi--.::osemia. None ol the other enzymes listed, .:nt. rv.ill give rise to the s1.mp1oms produced, .-.rrLr. cataract formation. A delect in gl,vcogen .-.' u'ould lead to reduced glycogen leve1s and
--:rssical galactosemia,

r.

:lpoglycemia. A defect in fructokinase leads -.-.surla (fructose ln the urine), but no ol'ert .:Ls of disease. The figure belorv lndicates the - lor galactose metabolism and the delects in . . .'nJ nonclasrr, al galactosemil.
n .

fhe answer is D: The high

NADH/NAD*

ratio impaired

Nonc assical galactosem a

Classicalgalactosemia

] Ga actose 1-phosphate uridylylkansferase :: -\ I+Glucosel-Phosphate ::-lOS

._.1

Grucose

6l:l1ose:,,"

----

(Lrver)

;,?yffI.J:.,

Glucose

-r*
-

answer is B: Reduced effectiveness of carboxylation '*<tions, leading to reduced glucose production' Raw'

-, -

..

:':arn a potent blnding partner to biotin, avidln, ', nile bound to biotin, blocks biotins particrpa' :.rrboxylalion reactions. This leads to reduced -.f pyruvate carboxylase, a necessary step in .:coneogenic path$'ays, thereby leading to a . -. :bilit,v of lhe ln er to properly marntain blood
ar-els. As oxaloacetate leve1s

drop due to lhe

gluconeogenesis. Ethanol oxrdation to acetlc acid (r'ia acetaldeh,vde) generates large amounts ol NADH. As liver glycogen stores har.e been depleted w'lthin 36 h ol the fast, gluconeogenesis is required to rnaintain blood glucose ievels. The major precursors for gluconeogenesis are glycerol, lactate, and amino acids (u'hlch give rise to pyrur.ate or TCA cycle precursors, u'hich generate oxaloacetate). Because of the hrgh NADHNAD* ratro (due to the ethanol metabolism), p;'r11'21. 4.ttined lor gluconeogenesis rs shunted to lactate in order to regenerate NAD* to al1or'v alcohol metabolism to contlnue. Similarl)., oxaloacelate is shunted lo malaie, also to regenerate NAD* lor ethanol meraboLrsm. Glycerol, rvhich is converted to glyssle|-3-phosphate, cannot go ro dihydroxyacetone phosphate due to the high NADH ievels in the lir.er. Thus, the hlgh NADHNAD* ralio drverts gluconeogenic precursors lrom entering gluconeogenesis, and the 1lr'er has trouble maintaining adequate blood glucose levels. Liver glycogen stores have been depleted r,vithin the first 36 h of the fast, but glycogen regulation is not affected by the NADH/NAD* ratio. Under conditions in lvhich the liver is exporting glucose (glucagon administration, for example), Iiver glycol,vsrs

84

Chapter 10

,Pyruvate

cH3cH2OH

,,t ,,
Gtucose

.;

,,

,' ,;

l\^NAD+ ILactate

I - rrnor-., l./-' +H+ (

7.., The answel is C: Aldotase. The disorder is herediranfructose intoierance, with a reduced abiiity to .or-rr..* fructose-1-phosphate to dihydroxyu..rorr. phosphate
and glyceraldehyde. The specific defect is in aldolase B. as much as g5%. This prob_ 1em is oniy evident when sucrose is introduced inro the

with its acrivir), reduced by

Ethanol
ADH

T
+

I\
I

NAD*

;'

runos

r-r*

" ,.

'.

^'
\.

-,-.

Oxaloacetate

diet, and fructose enters the lir.er. The accumulation

cH3

c:
I

.-O
H

..i

t\^NADi IMalate Dihydroxyacetone phosphate

l-- r.rnou r +Hl

Acetaldehyde
ALDH

mia Elimination of liuctose from the diet will reverse the symptoms. Galactokinase is needed for galactose metabolism; since the patient digests milk normally
galactokinase activity is not altered. Similarly glucose metabolism is not adyersely alfected (milk contains lactose, which is spltt into glucose and galactose), indi_ caring rhar herokinasc and glucokinrr. u,.,rr iries are normal. The defecr in aldolase B will hinder glycoly_ sis. but the lrr.er also contains aldolase C activity (thls isozl.ms u.tll not splir frucrose_1_phosphate), which

of fructose-1-phosphate, due to the reduced aldolase actl\drl, leads to a constellarion of physrological prob_ lems resulting in nausea, vomiting, urrd hypogly.e_

T
+

l\

NAD*

runon

H*

/ro cH3c\o
Acetate

l.
+

[.ii:'
,oo.

Glycerol 3-phosphate

1
I

Glycerol

A defi.ciency in fructokinase will lead to an accumula_ tion of frucrose (nor frucrose-1-phosphate), whlch is released into the urine (fructosuria), but does not lead
to the physiological symproms exhibited by the patlent. The fructose parhway (indicating the reactlon catalyzed

enables glucose metabolism to be very close to normal.

is inhibited by covalent modification of key regulatory enzymes, nor rhe NADI-VNAD* rario. These pathways are indicated in the figure above.

by aldolase B), and irs relationship to glycolysis, is

shor,vn beloui

Fructose )Frucrokinase ADP<'/I Fructose rlpnosphate

L t

Glucose ATP-I

ArP{

lo,oo,u""

t+

\ ^"-^DP4r+ \ / Glucoseo- <----+ Glucoselphosphate phosphate Frr.tlr"

o-

rc^u^r anc )Hexokinase f

Glycogen \

B
'--'--"'"':Hlv'Hrlqre

Glyceraldehyde ATp--l ,
) Trjose kinr.o ADP<'/|
Glyceraldehyde

}->

oir,vo.o*yacetone-phosphate

' !
I

I ruutvoe r'e-pr ruupllate Fructose 1,6_phosphate tAtdotase B ltiver) y'Aldolase A (muscle)

on:tof'" ( | \/

I l.

3-phosphate a Dihydroxy -,,'\ *-Glyceraldehyde 3_ """1nu_ phosphate _..v phosphate

+ +
Lactate

t'.^

<-----*

+
Pyruvate
I

N+ratty

acios

TCA cycle

Answer

Gl1'colysisandGluconeogenesrs 85
-he answer is

(:

-,:r has a BNII in the unhealthy range (15.7), incli:. inadequate nulrient uptake. Since her nutr.ienl
,..: is poor, most ol the glucose present rn her blood
as

GIycerol, Iactate, and

glutamine.

T1-re

is a mutation in pancreatic glucokrnase. lvlODY3 rs a rrl-itation in the trzlnscriptron factor HNFl-q, $.hiLe
N{ODY4 contains a mutation rn insulin promoter lactor 1 MODY5 rs a mLltalion in another transcription factor, HNFl-8. \,'IODY6 is a mur.rrion rn nerirogenrc difleren, tiation lactor 1 MODY is not insulin resrstance. ThereIore, all the other aspe cts of rnsulin resistance syndrome

: :::-r-ed lrom gluconeogenesis,

r
.rr.rSl

her glycogen stores

, -

. I
-

likeiy depleted. The substrates fbr gluconeo::,i are iactate (derlved lrom red bloocl ce11 meta:.1 i. g1i'cero1 (lrom trigll,ceride degraclation). and
:-

-. ;rcids derived lrom protein muscle degradation. ,,.lrine is a giucogenic amino acrd, anci is :r1so use d

:,i-rsport nitrogen groups lrom the muscle to the liver .., e excretion via the urea cy'c1e. Leucrne rs a slrictiv . .:rLic amino acid (gir-ing rise onl,v to acetyl-CoA), ' -'. .-:ucine carbons cannot be r,rsecl to make glucose . -iconeogenesis. Fatty acids are also stricLl,v keto- -. and cannot be used lor glucose producLion (latL;, . -.. qir-e nse to acetyi-CoA, r,vhich cannot be used Lo --.i.e net glucose). L1-srr.re is also a strictll'ketogcnie .- acid, and cannot be used lor glucose production. . -: clegradation gnres nse to bilLrubin, r,r,hich cannol ,.riher degraded, and none of the carbons ol 1-reme tr: used for glucose production.

are not presenl (obesrLy, hypertension, and hlperirrglyceridemra). Srnce \{ODY is autosomal dominanL, it can be traced through Lhe lamil; tree . It r.r,as thought at one time thaL the patient hacl to be Vou11g to presenl rvith tl'us disorder, but patients up to age 50 h:rve been

reported.
ce11

lt is not

t.vpe

diabetes mellitus as no islet

antibodles are present. Glucokinase is acting as a glucose sensor for the B-ceil. A mutated, less scnsitivc sensor leads to mildl1. eler.ated blood glucose levels.

t1

The answer is C: Salivary

" :,
'
,

- ... natural inhibitor of salivary

amylase.

Starch blockers conam1,1ase.

w'hich r.r.iil
am1'1ase

,,

-.. starch digestion in the mouth. The other

The answer is C: Galactokinase. The chrld has nonclassical gaiactosemia, a delect in galactokinase . \\'ith Lhis disorder. ga1:rctose .rannot be accumulaLed r,viLhrn cells, and so Lt accumulates in the blood. spilling or.er to the urine. Because ol rts hrgh level, the galactose can enter the e1-e and be reduced to gaiactitol b1'aldose reducrase. Lrapping Lhe galactitol r.vithin the eye. As galactitol accumulates, an osmotjc imbaiance is created, leading to caiara.rt lormatiolr. Hor,ver.er, slnce galactose1-phosphate is nLrt accumulatir-rg (as occurs in classicaL galactosemia, a deiect in galactose- 1-phosphate urid,vl1,l transferase), the other ellects seen u.rih classical galactosemra (hy.poglycemia and neuroiogical de ficit) do not

Jiqests starch, pancreatic amylase , u,ou1d onl)' be .' rrLed by the starch blocker if the inhibitor would
-i, e the conditions ol the acidic stomach r,r.,rth. recoming denatured. There is no gastric amylase.

:s:ina1 enteropeptldase rc[i\'.ltcs tr1-psrnogen, r,vhrch . :quired lor proteln digestron, not stalch digestron. : ,:::reatic lipase is lequired for dietary tnglycericle - .:strLrn, and is not active toward starch.

The answer is A: Pancreatic glucokinase. The boy -,. der-eloped MODY (maturitl. onset diabetes ol the
. -r::q), and one varianl of MODY is a mutated glucoki-

inheritable disorder) such that the K, lor gluinsuhn release only occurs r'vhen --:l . :erglycemia is present. Both an increase in AfP and l,:,DPH are required lor the pancreatic B-ceil to release

,::.

,\an

has increased, and

.-lhn. When pancreatic glucokinase has an increased ior glucose, ATP ievels can only increase at greater
,,n

occur. The sugar that is accumulatrng in the urine is galactose, u,hich contains an aldehyde, r,vhich generates a positive response ln a reducing test. A defect rn fructokinase leads to fruciosuria, a benign condition (fructose is not a substrate lor aldose reductase, rs it is a ketose and not an aldose). A delect in hexokinase r,vouid lead to eler.ated glucose levels, and can leacl tir sorbitol production in the lens ol the eye, but the urinc reducing sligar test was negative lor glucose. A delect in aldolase u,ould lead to the intracellular accumulation of metabolrtes, but not 3 grert increese rn circillating galactose. Refer to the figure in the ansrver to qllestion 3 of this chapter for the pathway of galactose metabolism and the enzyme delects in both classical ancl nonclassical galactosernia.

normal lel.els of glucose. Thus, moderate hyperglyia is not sr-lfficient to induce insulin release. As insulin -::l. '.-:ase occurs from the pancreas, liver, muscle, or inles..:-,-rl hexokinase wr11 not aflect the process. The pancreas *--.s not express hexokinase, oniy glucokinase. NIODY : :. monogenetic autosomal dominant disease of insulln ::Jretion. There are at least six amino acid substitutions , rr!r\\Tr in a number of diflerent proteins. MODY1 is a :r'tatlon in the transcription factor HNF4-cx- MODY2

t2

The answer

is E: Phosphoglycerate kinase. In

g1u-

coneogenesis, phosphoglycerate kinase catali,'zes the

phosphorylation of 3-phosphoglycerate to 1 ,3-brsphosphoglycerate, a step which requires AfP The other two steps requiring a high-energy phosphate bond in the
lase and

conr.ersion of pyruvate to glucose are pyru\rate carboxyphosphoenolpyruvate carboxykinase. Fructose-

1,6-brsphosphatase

and

glucose-6-phosphatase are

86

Chapter enz)Tnes

l0
that remove phosphates from
substrates, isomerase catalyzes the conversion of dihydroxyacetone

releasing the phosphates as inorganic phosphate. They do not require, nor generate, AfP Pyruvate kinase rs not utihzed for gluconeogenesis, and triose phosphate

phosphate and glyceraldehyde-3-phosphate, without the involvement of a high-energy phosphate bond.

These are shown

in the pathway

be1ow.

Glucose

Glucose 6-phosphate

I
Fructose 6-phosphate

lfr-:r"1
I

6-bisp hosp hatase

Fructose 1,6-bisphosphate

Dihydroxyacetone-P
I

Glyceraldehyde-3-P

Glycerol -..+Glycerol-3-P

t
t I

REOUIBES ATP (to convert 3-phosphoglycerate to 1,3 bisphosphoglycerate)

t
^/ REOUIBES GTP

7z

Phosohoenolpyruvate

nhosOnoenolpyruvate carboxykinase

Oxaloacetate

t
REQUIRES ATP

Amino acids Alanine

:xil:*,,,,1,*"(
Lactate

13

The answer is D: Pyruvate carboxylase. The bodys major energy source for gluconeogenesis is fatty acids, which are oxidized to acetyl-CoA, at which point acetylCoA enters the TCA cycle to produce AfP Acetyl-CoA actlvates pyruvate carboxylase (and inhibits pyruvate dehydrogenase), a key gluconeogenrc enzyme. Acetyl, CoA does not regulate any of the other enzymes listed as potential answers (PEPCK is transcrlptionally reguiated by CREB; Fructose- 1 ,6-bisphosphatase is inhibited
by fructose-2,6-blsphosphate; glucose-6-phospharase is regulated by a regulatory protein; and pyruvate klnase has both allosteric and covalent controls rn the liver, but none involve acetyl-CoA).

induced phosphorylation

of

PFK-2, which activares

PFK-2 phosphatase actir-ity, which convefts fiuctose-2,

6-bisphosphate to fructose-6-phosphate. Pyruvate kinase actrvity, in the 1iver, is also reduced by phosphorylation

by protein klnase A (which rs activated by glucagon). As blood glucose ievels have dropped during the fast, and the liver is exporting glucose, the concentration of glucose in the hepatocyte is not sufflcient for glucokinase (which has a high K_) to phosphorylate glucose. Glucokinase is not regulated by phosphorylation.

l5

t4

E. Under the condirions of a,l8-h fast, the Iiver is exporting giucose, and glycolysrs will be inhibited. PFK-1 actit ity rs reduced due to a reduction of fiuctose2,6-bisphosphate levels, brought about by glucagonThe answer is

The answer is A. Upon insulin release, the cAMP phosphodiesterase is actlvated, reducing cAMP leve1s in the Iiver, thereby leading to inactivation of protein kinase A. In addition, protern phosphatase t has become active and dephosphoryiates the en4rrnes that were phosphorylated by protein kinase A. Therefore, PFK-2 is not phosphory1ated, which leads to an active kinase actir,rty and an

.t-a

t'/
lr'*-"'

:-:.

': - ,.::-. .-h-'ice s -\ rhrough C. comblned r'rirh the - - - -' :::lns. rrnh choice A can be correcl)'
-rr,e answer is D:

--- :la '1:'= -'.-'.-1 r FFK-- .:'ils\ie:: ,.,-,4-aaa iil:n FiK-l 'tclnilr' rltr\\ trilh _. _t_-^- *-JLrrrlL\r'(LLJ yrv :ra CtrrllCl'. Llt)uIrIr ti Lfr raLL lnsulin stimulates pre, ,:.:l : :::.is:orters in the mr-rscLe to fuse uith the -:ll- :::.1e. :herebr enhancing glucose transport

::-i

K- -\TF:': gradieni ,.'.h,ch p.,r,-rps sodium out ol the ce1l tthree iltrlrls uri sodium.) in exchange for potassium (nr'-o aton-rs ol

:...r:'.:

ihal is esr-tlll=hed lli thc \'r '

-::-'.t

: -iI-

"'

'f':

2.phosphoglycerate. Fluonde inhibits .n:\'me enolase,lvhich cata11'zes the dehy- . -,. -.--:i.rsphogll'ceratetophosphoenolpyru\rate'

under -':-.isphoglr-cerate accumulates

these

-t answer is E: Na*, l(* ATPase. N{ost monosaccha-: . r: ':,lnsported. wllh sodium from the intestrnri

(outpotassium). This crea[es both a sodium gradrent (outside concentration higher) and a charge gradrent slde posttive as compared to inside the ce1l) across the plasma membrane. Due to these gradients, the entr,v of ,odt.,- into the cell is energericalll' lavorable, and the monosaccharide piggybacks u'ith the sodium for transporl into rhe cel1. The Na /Hn exchanger is not operative in intestinal eptthelial cel1s, and none of the oih., .rt.y-es (glucose-6-phosphate dehydrogenase' hexokinase. and chlorlde transporter) u'i11 create the necessarv sodium gradlent for monosaccharide transport. Carbohl'drate transport into the enterocl'tes rs outhned in the figure belor'li

Lumen Na* Mucosal side

catactose

J lL
(

Brush border

--r.-r

lntestinal epithelium

Serosal side

ui
The answer
-..

Nu.-orr.ose

cotranspofiers i]?,

Facilitated glucose

transpoders

.]

*".'^*-ATPase

is A:

Enhanced production

of

fructose'2,

t9

The answer

6-bisphosphate. When phosphorylated,-heart PFK-2

glucose and insulin levels was symptomatic' This pattent measured while he

is D: Blood

Phosphorylation of heart PFK-2 can be protein kinase ,, ---.-,mplished through the AMP-activated .. hen the heart is having trouble generating energy) -: in response to insulin (lnd'icating that high levels of ofheart :-:cose are avaiiable for use) Phosphorylation .:X-Z do"t not affect its transcripiion or turnover rate' .:cl also does not affect the degradation of fructose-l'
,, -: o[ the heart.

. ;tir.ated to produce more fructose-2, 6-bisphosphate glycolytic .-- .timulate heart PFK-l and to increase the

probably has an insulinoma that releases insuhn inapp.op.rui.ty at any blood glucose level, which u'ould i.rd ,o hypoglycemra. The released insulln would stim-

.rlot. g1r.ot. uptake into rhe peripheral

tissues- (muscle

^-brsphosphate.

blood glucose u,-ra fa1), and if the patient had not eaten, 1a11. The lnsulin is also inhibiting 1eve1s wou1d" rapidiy the liver from producing glucose, either from glycogen degradation or gluconeogenesis, which only comand resultant io.,r-tai rhe problem. The hypoglycemra for all ol his symptoms Lpinephrine release account

88

Chapter 10 To diagnose an insulinoma, a 1ow blood giucose 1eve1 and an lnapproprlately high insulin 1eve1 during ryrnptoms must be documented. A fasting blood glucose and HbAlc could be perfectly normal and do not help in making the diagnosis, as insulinomas do not necessari1y constantly secrete insulin; they do so episodically
The presence of islet ce11 antibodies helps diagnose t)?e 1 diabetes mellitus, but not an insullnoma. A noncontrast CT scan might be used to locate the position of the insulinoma, but it is a very poor test even for iocation The answer is B: A high

,f, for glucokinase.

,n. ,t""rl

;;il:'"'fi ,3::".:ffi';JTi,:?:,T,'lI;ff

:llH:l
t I

m[riusir*[::ri#:::' n*I
levels after a meal. Under normal, fasting conditions, the concentration of blood glucose is lower than the K-

and would only be attempted once the diagnosis was

confi.rmed.

Chapter

TCA Cycle and Oxidative Phosphorylation

'",rlrcr- contains questions on tlse TCA cydc

(A) Ethanol is conr.erted to acetone, and the carbons

are lost durlng exhalation

.r.

ri

r e pha

sphorylcttion, including

que

stlons

vith other aspects of metabolistxl. MetG-j.rses affecting a,spects of the TCA rycle .,lltrrtle phosphorylation arc also covered
": ;l

(B) Ethanoi is lost directl1. in the urine (C) Ethanol cannot en[er the lir-er, u,'here gluconeogenesis predominantly occurs

,:tll)ter. _

(D) Ethanols carbons

are iost as carbon droxlde before

a

a gluconeogenic precursor can be generated

L
'c,rs-*

TIONS
single best answer.

(E) Ethanol is converted to lysine, lr.hich is strictly

ketogenic amino acrd

::r

il - - :,.nic alcoholic, while out on a binge, became verl' .,-sed and forgetfui. The police found the man and - -:nr hrm to the emergency department. Upon exam-,..r:r. he displayed nystagmus and ataxia. Which
----:ie
rs
:

displaying reduced activity in his brain under

-. -h ceraidehyde-3-phosphate dehydrogenase - .socitrate dehydrogenase - u-ketoglutarate dehydrogenase - Succinate dehydrogenase

\lalate dehydrogenase

..:

conditions?

A family that had previously had a nervborn bo1, dle ol a metabolic disease has just gi-,,en blrth io another bo1,, smail for gestational age, and with 1ow Apgar scores. The chrld dlsplayed spasms a few hours after birth. Blood analysis indicated extremely high 1eve1s of lactrc acid. Analysrs of cerebrospinal fluid showed eler,ated lac[ate
and pyruvate. Hyperalaninemia lr,as also observed. The child died within 5 days of birth. The blochemical defect in this child is most like1y which of the follou.ing?

(A) (B) (C) (D)

The

El subunit of pyruvate dehydrogenase

The E2 subunit of pyruvate dehydrogenase The E3 subunit of pyruvate dehydrogenase

Citrate slmthase

:: energy

yield from the complete oxidation of acetyl-

(E) Malate dehydrogenase

-,

., to carbon dioxide is which of the following in terms

:. :rgh-energy bonds formed?

3-month-oid

girl

developed

lactlc

acidemia.

.-6 :8 :10 t12

t,l

Blood analysis also indlcated elevated levels of pyruvate, cr-ketoglutarate, and branched-chain amino acids. A urinalysls shor,ved eler.,ated levels of lactate. pyruvate, o-hydroxyisovalerate, o-ketoglutarate, and o-hydroxybutyrate. A likely mutation in which ol the following proteins would lead to thls clinlcal finding?

:-eanol ingestion is incapable of supplying carbons --r gluconeogenesis. Thls rs due to which of the fo1:-,irng?

(A) (B) (C) (D)

The

El subunit of pyruvate dehydrogenase

The E2 subunit of pyruvate dehydrogenase The E3 subunit of pyruvate dehydrogenase

Citrate slmthase

(E) Malate dehydrogenase

89

E=:

E-::. . : ---' E :i:. ,-r

F::.:;:.-=::.=:..:a':a

90

Chapter 11
A human genetlcist is studying two different families. In one famiiy, all of the children of a mildiy affected mother display myoclonic epilepsy, developmental dlsplay, and abnormal muscle blopsy (ragged red fibers). In the other famlly, the three children of an affected woman endure energy yield for the complete oxidation of citrate to six carbon dioxides and water is which of the following? (A) 1 5 .0 moles of AfP per mole of citrate (B) 17.5 moles of AfP per mole of citrate

strokellke episodes and a mitochondrial myopathy The common link between these two diseases is r,vhich
of the following? (A) Mutations in (B) Mutations in (C) Mutations in (D) Mutations in (E) Mutations in pyruvate dehydrogenase complex cytoplasmic IRNA mitochonddal IRNA
malate dehydrogenase pyrur:ate carboxylase

(C) 20.0 moles of ATP per mole of citrate (D) 22.5 moles of ATP per mole of citrate (E) 25.0 moles of AfP per mole of citrale

tt

You have been following a patlent for several years, who

has recently become clinically depressed, and is eating very little and drinking alcohol r.ery heavily He presents

to you one day with nottceable swelling of the lower Iegs, lncreased heart rate, lung congeslion, and complaints of shortness of breath with virtually any acti\1t)I These sl.mptoms have come about due to which of the following? (A) Lack of energy [o the nervous system due to niacin deficiency (B) Heart has trouble generating energy due to niacin deficiency (C) Lack ofenergy to the ner\.ous system due to B, deflciency

A toddler has been diagnosed with a mild case of Leigh slndrome. One possible treatment is which of the folIowrng?

(A) Increased carbohydrate diet (B) Additional Bu in the diet (C) Decreased lipoamlde in the diet (D) Addltional thlamine in the diet (E) Decreased fat diet

(D) Lack ol energy to the heart due to B, defi.ciency

A patrent was diagnosed with a mitochondriai DNA

(E) Lack

i activity This patient would have difflcultles in which of the followlng

mutation that led to reduced complex
electron transfers?

of TCA cycle activity in the kidneys, leading to excessi\ e water retenlion

tz An S-month-old girl was taken to the

emergency

(A) Succinate to complex lll (B) Cytochrome c to complex IV (C) Coenzyme Q to complex IIl (D) Malate to coenzyme Q (E) Coenzyme Q to oxygen A pair of farm workers in Mexico was sprayng
pestr-

department due to the onset of sudden seizures. The chrld had brittle hair, wtth some bald spots, and skrn rashes. An ophthalmologist noted optic atrophy. Urinalysrs showed slightly elevated ketones and the presence of other organic acrds (such as propionate and lactate). Tieatment of this child with which of the following can
successfully alleviate the problems?

cide on crops when they both developed the following severe syrnptoms: hear'ry, Iabored breathing, significantly elevated temperature, and loss of consciousness. The pesticide contained an agent that interfered with oxidative phosphorylation, which most closely resembled whlch of the following known inhlbltors?

(A) Thiamine (B) Niacin (C) Riboliavin (D) Carnitine (E) Biotin
13

(A) Oligomycin (B) Atractyloside (C) Cyanide (D) Rotenone (E) Dinitrophenol
10

The refi11ing of TCA cycle intermedlates is frequently dependant upon which of the following cofactors?

crazed friend of yours has gone on an orange juice, fi.sh, and vitamin pill diet. He tel1s you that the citric acid, slnce it is a component of the TCA cycle, is always recycled and does not count toward his calorlc total each day You disagree, and inform him that citrate can, in addition to har.rng its carbons stored as glycogen or fat for later use, produce energy for hrs daliy metabolic needs. The

(A) (B) (C) (D) (E)

Niacrn Riboflavin Carnitine Pyridoxal phosphate

Lipoate

t4 The concentration of TCA

cycle intermedlates can be reduced under certain conditions. Consider a patient who inltiates taking barbrturates. During the initiai phase of his taking this drug, which TCA cycle intermediate is reduced in concentration?

TCA Cycle and Oxrdatrve Phosphorllatrc.n 91

l7 An inactivating mutation ln \ ''hich of the

-..-Llll-2lr - - -:-'.'L-L,il^

foLlou'rn:

enz),mes would lead to laclic acid accumulation rn

thr

liver?

-i::.ia

- - .;:elate
rllilmur:lil:r. m: - i and 16 are based on the following graph of oryrilr '::ir - llion by carefully washed mitochondria as a funcrtrrmrr ::-. {TP. ADP, inorganic phosphate, and oxygen are flmr-qffi:r ::: no oxidizable substrates. Once a compound is ilflililufl: :: mixture, it is not removed, nor is the length of the rllrnnLurf*-:r:f::. suf0cient to use up all of the compounds added to

(A) (B) (C) (D) (E)

18

Glucokinase Phosphofructokinase- 1 C)toplasmic malate dehydrogenase Pl.rur.ate kinase

Glycerol-3-phosphate dehydrogenase

researcher was stud,ving oxidatjye phosphorl.latron

lltrln n:

I '::.

ndriOn.

1=

Pyruvate

ancl isolated mitochondria. ATB ADB inorganic phosphate, lactate, lactate dehl.drogenase, and ox)rgen were introduced to the suspension, and he $'as able to demonstrate ATP productlon rvithin the mitochondna. The researcher then added oligomycin to the mlxture, rvhjch stopped oxygen uptake. This occurred due to rvhich ol the follor,vrng?

in a suspension of carefully rvashed

2 = Succinate 3 = Oligomycin 4 = Cyanide

(A) (B) (C) (D) (E) t9

lnhibrtion ol compler I lnhibrtion ol cornplex I1 lnhibition ol complex Ii1 lnhibition of complex IV lnhlbrtron ol the proton translocating ArPase

A ner,vborn displa,vs lethargy and crying episodes. Blood ana11.srs indicates lactic acidosis and hyperalaninemia.

In order to distingulsh
Time

betr,veen

a pyruvate dehydro-

genase complex deficiency and a pyruvate carbo$4ase deficrenc1,, one can measure u,htch of the fo11ou ing in

the blood?
.,:: compound was added at the point indrcated -\ntimycin A
as A?

: -

\tractyloside
Rotenone

(A) Fasting blood glucose (B) Alanine aminotransferase acti\,ity (C) Free latty aclds levels when fasting (D) lnsulin 1eve1s rvhen fasting (E) Glucagon levels u,hen lastrng
20 Your obese patient has tr-pe 2 diabetes mellitus and you har.e started him on metformin. One of the possible complications ol metformin therap; rs lactic acidosis. Why.is this a concern u.ith metlormin therapy?

Dinitrophenol

- LacLate
'.:.Lat

compound was added at the point indicated as B?

-, l't :I

Antimycin A
Atractyloside

.^' Rotenone
Dinitrophenol
Lactate

(A) (B) (C) (D) (E)

Metlormin reduces lnsuhn resistance

lr"letformrn blocks hepatic gluconeogenesis N,letformrn blocks the TCA cycle Nletformrn inhibits glycolysis

Metlormrn rnhibits dietary protein absorption

92

Chapter 11
molecuies of NADH are produced, along with one mtecule of and one substrate_levJ phosphorl.l;._ .FADH, tion resulttng in the generation of GTp A; each NADF,

ANSWERS
,1r,,, The answer is (: a-ketoglutarate dehydrogenase. The alcohohc has become deficient in vrtamln S,. thiamine, which rs converted to thiamine pyrophosphate for use as a coenzyrne. One of the symptoms of B, deficiency

oxidation to generate energy

dehydrogenase, and u-ketoglurarate dehydrog#ase. By reducing the activity of the latter two enz),tnes, glucosl
1s

lumber of enzymes, including transketolase, pyruvate

.3

is neurological, due to insufficient energy generation r.vithin the nervous system. B, is require-cl for a smal1

ls 10 (7.5 from NADH, I.5 from FADH, and 1 fior:GTP). Thrs rs shown in the flgure below.
The answer is D: Ethanols carbons are lost as car. bon dioxide before a gluconeogenic precursor can be

can gir.'e rise ro 2.5 Afll and each FADH, to 1.5 ATp r: oxidative phosphorylarion, the net yield of high_energ. bonds from one acetyl-CoA being oxidized by-the cvci.

vous system suffers because of it.

impaired, and the

ner_

carbon dioxide, and oxaloacetate is regenerated, three

The answer is C: 10. When acet1,1_CoA enrers rhe TCA cycle, and is conr.erted ro two molecules of

erate energy, and two carbons are lost for each Lurn othe cycle as COr. Thus, ethanol cannot provide carbons lor the net s).r-rrhesis of glucose. Ethanol is not converred

is converred ro aceraldehyde, whicl: is further oxldtzed to acetic acid and is then activated rc acetyl-CoA. The acetyl-CoA enters the TCA cycle ro gen_

generated. Ethanol

crr.C'3icon

COO-

Acetyl CoA
I

3=o
***"#71::Z

coASH

,/
Ho-3n
?oo-

oxaroacetate
+

/citrate"vntnu"\ Hzo [oo]:5-';"" ?rr3H,

F--*or^
*^r-

coo-

/\ 9Hr' I

H.

--........
'..

i:\o.".,,,"" coocitrate

\f.

clz
I I

i I tx (

H_C_COOEt_ectron_...

t}:=l'.

'..

Ho-c-H
COO-

Hd tt

Oxidative phosphorytation

^r

\x

\!t"'", \ t'.
I '"
-.:

Nnn*rl
llsocitrale COO- dehydrogenase
I

Fumarate

coo-

CH.': I

HzO Oz,i

-J

FAD(2H)

i NADH
+H* -+'

,'

"r" ir,
I

NAn+

,/

?=o

succinate

/ iy $ succrnate ^:^ +P, irffiffi GrP

Coo-

'\>\_

CH2

J=o
r

ioASH

fid#rH::,:

scoA
Succinyl CoA

Answer 2: The Krebs tricarboxylic acid cycle.

TCA Cycie and Oxidative phosphon,iarron

-.: ::.r is it dlrectly lost in the urine. Ethanol -' - . ' -.ridrzed 1n the 1iver, and its carbons can_

93
a:t

': ,:: j ror the biosynthesis which is - ,,:t-no acid for humans.of lysine, oxidatron an Ethanol is -,_ - :he figure below.
cH3 CH2OH

Ethanol
I

l/_ NAD_ ADH t\ ITNADH+H* v

T

would not lead to severe lactic acidosrs andwould nor be male-specific disorders. As an example, the three subunits of cr-ketogiutarate dehydrog..rrr. are shown
be1ow.

that catalyze oxidatir.e decarboxylation reacrions. and other metabohres would also be accumulatlng. Defects in cirrare synrhase and malate dehydrogenai

E3 deficlency would affect more than pyruvare metab_ olism, as this suhunit is sharecl with other enz)-tnes

from rhar of an E2 or E3 deficiency In additron.

The answer

cH.c':o
I

drogenase. The child

Acetaldehyde

l'ALDH
V
I

t'tRo'
runoH

l\
T

of
*
l.r.

c.,ketogiutarate dehydrogenase, leadrng to the tuildup

drogenase, leadlng to a buildup of lacraieand p),ruvaie,

oxidative decarboxylation reacrions (pymvate dehy, u-ketogiurarare; and branched_cilin cr_ketoacid to a burldup of many of rhe -

is (: The E3 subunit of pyruvate dehy. is defective in a varlety oI

dehydrogenase, leading

zro CHsC_o_
Acetate Ethanol metabolism.

-le answer is A: The El subunit of pyruvate dehydro!rase, Lactic acidosis can result from a defect rn an : -- -iu3 rhat metabolizes pyruvate (primarlly pyruvare
:ehvdrogenase complex conslsis of three ma.;or .,..,.-.':rc subunits, destgnated Ei, E2, and E3.

dihydrollpoyl dehydrogenase activity, is common among these enz)..mes. Thus, a muration in E3 would render all of these enzyrnes inoperable, leading to a buildup of the.cx-ketoacid precursors. Defects in ciirate s)nthase or malare dehydrogenase would nor leacl to the buildup of these cr-ketoacids. The answer
eases. Family

other metabolites). Enzlmes, whtch catalyze oxiclative decarboxylatron reactions, contatn three catalytic sub_ units, E1, E2, and E3 (see the figure in rhe answer ro the previous quesrion). E3 subunit, which contains the

-r,.

-iogeoi1Se and pyruvate carboxylase). The pyru_

is rhe one that binds thiamine pyrophos_ --:.: and catalyzes the decarboxylation of-pyruvate. - ,: =ene for the El subunit is on the X chromosome, . . ::tects in this subunit are inherlted as X_hnked -.::ases, which primariiy affects males. Since this is .. second male child to have these symptoms, it is ...:,r- that the mother is a carrler for this disease The ::,.rn of inheritance distinguishes thls diagnosrs

, .'.:unlr

The

Both famtlies are suffering from mitochondrial

is (: Mutations in mitochondrial

tRllA.
dis_

tion in tRNALv., whereas MELAS has a mutation in a tRNAieu gene. In both cases, the IRNA mutations inter_ fere. with protein synthesis within the mirochondna,
leading to
a

ragged red fibers) while family 2 has MEf_ab (rrrirochondrial myopathy, encephalopathy, lactic aciclosis, and stroke). Both disorders are due to mutations in a mitochondnally encoded rRNA. MERRF is a mura_

has MERRF (myoclonic epilepsy with

reduction of functionai proteins necessar\-

ot-t
I

FI* C -TPP

.:.

- :- Lhree different subunits are required : :eacrion, EI (a,keto acid decarboxv-i (transacylase), and f; (dihydrolipoyl - _:--genase). Tpp to the .. . :rrophosphate.refers refers cofactoithi_ Lip
to the cofac_
-: ---c acrd.

of g_keto acid dehy-1-:_ie complexes. R represents the por- -:re cx-keto acid that begins wltn *re B

i:: *: Mechanism

Hl

tl

FAD (2H)

Dihydrolipoyl DH
E5
t

NAD+

CO,
D

t s
,rv

cl-Keto

acid DH

t,i

i6) E1 6i
'.
V.

F\ .,,--.',s. .1.

r.

,-r.*---,

o-Keto
TPP

':

|"

trans Ac '.

l__"_-"_j

COO-:

acid DH

.,-.

a-Keto acid

HS S-C-R

./ \

LiP

o
ll

94

Chapter 11

aspects of oxidative phosphorylarion. These disorders are not due to mutations in nuclear encoded genes (which eliminates all ol the other
answers).

for various

The answer is E: Dinitrophenol. The key is the elevarion in temperature. Dtnitrophenol is an uncoupler of oxi_ dation and phosphory4ation in Lhar uncouplers desrroy

the proton gradient across the membrane (thereb1, rnhibrting the synrhesis of ATp) wrthout blocking the

The answer is D: Additional thiamine in the

diet. Leigh

syndrome can result from a deflciency of pyrur.,ate

dehydrogenase (PDH) acrir.ir)., leading to lactic acidosis. In some cases, the enzyme has a reduced alfinity

Ior thiamrne pyrophosphate, a required cofactor for the enzyme. Adding thiamlne ro rhe diet may overcome this deficiency. by raising the concenrratjon of thiamine pyrophosphare such thar it will bincl to the altered enzyme. Increasing the carbohydrate rn the dlet will make the disease worse, as more pyruvate would be generated due to the increase in the glycolytic rate. Vitamin Bu does nor play a role in glycolysis or the PDH reaction. Lipoamide is a required cofactor for the PDH reactton, so reducing lipoamide would have an adverse elfect on the activrty o[ PDH. Decreasing the fat content of the diet may be harmfu'l, parricularly if the calorles are replaced as
carbohydrate.

in the mitochondrial matrix, electron florv r,vi11 stop due to the inability ro synthesize ATp (normal
ADP couphng). Oligomycln works in a srmilar mechanism

transfer of electrons through the electron transfer chain to oxygen. The energy that should have been generated rn the lorm of a proton gradient is lost as heat, which eler.ates the body temperature of the affected r,vorkers. Electron flou, is also enhanced in the presence of an uncoupier, so additional oxygen is required to ailow the chain to conrinue (hence the heavy breathing). The other agents added would have stopped electron transfer tota11y, whtch wouid not a11ow for an increase in temperature, and wouid actually decrease the rate ol breathing (since oxygen is no longer required for the nonfuncttoning electron transfer chain) Atractyioside inhibits the ATP/ADP exchanger, and once there is no

in that it blocks the ATP synthase, prevenring ATp synthesis, and, due to coupling, elecrron transfer through the chain. Rotenone blocks complex I transfer to coenzyme Q, which significantly reduces electron flow, and will not lead to an increase in temperarure.

The answer is D: Malate to coenzyme Q. Complex i accepts elecrrons from NADH, and will transfer them to coenzyme Q. Malate dehydrogenase will convert malate to oxaloacetate, generating NADH ln the process. The NADH wili then donare elecrrons to complex I to initiate electron transfer. Succinate donates electrons at complex Il (r,ra succinate dehydrogenase, a componenr ol complex 11), which donates to coenzyme e, thereby bypassing complex I Cytochrome c transfers electrons from complex IIi to complex IV Once electrons are carried by coenzyme Q, complex 1is no longer required for electron transfer to oxygen. These transfers are outlined in the figure below

t0

The answer is D: 22.5 moles of AIp per mole of citrate. The followlng sreps (see the figure on page 95) are

required for the complete oxidation of cirrate to carbon dioxide and water. First, citrate goes to isocitrate. which goes to o-ketoglutarate (thls last step generares carbon dioxide and NADH, which can grve rise ro 2 5 ATP). The s-keroglurarare is furrher oxldized ro succinyl-CoA, plus carbon dloxide and NADH (this is the
second carbon released as CO,, and another 2 5 ATp)

Succin; I-CoA is converted ro succinale. generar ing a GTP (at this point, five high-energy bonds have been

created, plus two carbons lost as carbon dioxide).

12

02 +

NADH dehydrogenase

2H+

H2O

Succinate ETF: Q Cytochrome b*c, dehydrogenase oxidoreductase complex

Answer 8: Electron flow through the electron-transport chain

Cytochrome c oxidase

Matrix

TCA goes to tumarate, with the generation ol another 1.5 ATP), fumarate is converled lo .:. and malate leaves the mitochor-rdna (r.ia the . .: -..sparlale shuttle) lor lurther reactions. Once : .\topiasm, the malate is oxidized to oxaloace, ..reratlng NADH (anoLher 2 5 ATP il the malate/ -. r:.-c- shr-rttle is usecl). At this point, citrate has - -..nYerted to cyLoplasmic oxaioacelate, u'ith tl-re : r-,'-rrrr1 ol te n high-energ;, bonds and the loss of -.ribons as carbon dioxide. The oxaloacetatc is -,rnlerled to phosphoenolpln-u'ate and carbon --. at the expense ol a high-energl' bond (GTP, :r:osphoenolpvruvate carbon-kinase reaclion). : -,-:h-energy bond is recovered in the nert step. :''ar. as PEP is conr,erted lo pyru\rate, generxtlng - I Thus, ai this poini rn our convcrsror., Litrate : :--:-re to p).r1l\rale , plus three CO., ri,ith a net yielc1 . : \TP (or high-energy bonds). The p1'n-rvate re.:': tl-re mitochondria and is oxidized to acet,vl-CoA

C1.c1e

and Oridatrve Phospliory-latrorr

9;

,i:-

11

of energy to the heart due to B, deficiency. The patient has thiamine cleficiencr. -:r-.The answer is D: Lack because of this, his heart ls having troubie generrtln:l sulficient energ), to elfectively, pump hrs blood (due t.. a reduction in the rale ol boLh pvruvate oxidation ar-id TCA oxiclatir-e steps). The resultant congesti\-e hearr failure leads io eclema in the lor'ver extremities, puLmonar1. edema, and inabiliLl Lo pirtiuil.lte in er-en miid exercise . The thiamir-Le deficrenc,v has resuited from the patienls 1.roor diet and the ellecL ol ethanol blocking thiamine absorption lrom the diet. The neryous system also sulfers lrom thjamine deficienc1,, in r,vhrch case, neuroLogical signs ol the deficiencl' u.or-rld be er,'ident. These are not ,vet obsen.ed irr this patienl. The s1.mptoms obsen.ed are not due to niacin cleficrencl. (lvhich are dementra, derrr-ratitis, and drarrhczr). The problem is also nol clue to insulftcient energ) lor the kidnev to appropriatell, fi1ter the blood

- ::rbon dloride. also generating NADH (another ' -TP). \\ihen thrs acet1.l-CoA goes around the TCA - :. t\\'o carbon dioxrde molecules are producecl, -. u'ith another ten high-energy bonds. The net
,,, is therelore six carbon dioxide molecules and : :iqh energl.bonds lor the complete oxrdation ol

t2

Biotin. The child has biotrnrdase defi, in a functional biotin defi,,icnur Bjotinidase is required to rernove cova1ent11. linl<ed
The answer is E:
cienc1., u,hrcl-i

results

biotin from proteins in our diet and from proteins that have turnecl o\-er \\-lthin the body An inabrlitl'to do tl:ris leads to a biotin defrciencl. (as most ingested brotin is

NAD+ NADH
Citrate
lsocitrate

NAD* NADH

\4

COz
Malate

)r

o-ketoglutarate

+
COz

Succinyl-CoA GDP GTP

tmito)

) - HOH

Fumarate

FADH2

t * ,5\

Succinate

FAD

Overall, then, there are

6 carbon dioxide generated

7 NADH (which yield 17.5 ATP) 2FADH? (which yields 3 ATP)

1 GTP 1 ATP

For a total of 22.5 moles of ATP per mole of citrate

Answer l0: The parhl,ay, required Ibr the compleLe oxidation of cttrate to carbon dioride and u.ater

96

Chaprer

11

linked to proteins). The hair and scalp problems have been attributed to an inability to s)'nthesize fatty aclds (as acetyl-CoA carboxylase is missing biotin). Since pyruvate carboxylase is also inoperative (due to the lack of biotin), gluconeogenesis is impaired, and ketone bodies will be sy,nthesized by the liver to compensate for reduced glucose production. Priopionyl-CoA carboxylase is also impaired, leading to the elevated levels ol propionic acid. Since gluconeogenesis is impatred, excess p)rruvate rrrl1 be converled to lactate since it cannot be converted to oxaloacetate. The optic atrophy may be due to an inability to s),nthesize fatty acids within the neurons or a iack of energy due to reduced gluconeogenesis.

,;4

(: Succinyl-CoA. Barbiturates are metabcItzedvia cytochrome P'150 enzymes, whlch are rnduce: by their substrates. The lnduction of slnthesis require. that heme be sl,nthesized, and the first step in hern.
The answer is

s)'nthesis requires succinyi-CoA and glycine and occurs

wrthin the mitochondrial matrlx (see the figure belou' Thus, succlnyl-CoA lel,els can drop in the matrix durin-. heme synthesis, and anaplerotic reactions are require; to keep the cycle going.

cooI

cHz
I

CHz
I

t3

The answer is D: Pyridoxal phosphate. Pyridoxal phosphate is required for the transamination of aspartate to oxaloacetate and glutamic acld to cr-ketoglutarate. Both the cx-keto acids are TCA cycle components, and

C=O
I

SCoA

Succinyl CoA
+
H2c
I

when thelr levels decrease, they can be replenished

through such a reaction. Niacrn, nboflavln, and lipoate are required for oxidative decarboxylation reaclions, but that reaction rype does noi lead to a refilling of TCA cycle intermedlates. Carnitine is required to transport acyl groups into the mitochondna and is not used io transport TCA cycle intermediates from the cytoplasm to the mitochondria. Biotln would be a correct answer (for the plruvate carboxylase reaction, to regenerate oxaloacetate from pyruvate), but lt was not offered as a choice. A typical transamination reaction is shown below. A

fiH3

cooGlycine

,., coAs-.y'\ co' V

syntnase

u,oro [
I\

cooI

CHz
I

CHr

C=O
I

t-

H2C

tiH3

6-Aminolevulinic acid
(5-ALA) The first step in heme biosynthesls.

coo+l H"N-C-H "t

CHz

cooC=o
I I

15,

CH,

coo-

Aspartate
cooC=o
I I

>H

tcoo-

The answer is (: Rotenone. At point l, an oxidizable substrate was added io the mixture as indicated in the figure (pyruvate), which is oxidized to form NADH. The

Oxaloacetate

cooCHz
I

NADH can add electrons to complex I to initiate electron flow across the chain. Since at point 2 the addltion of succinate ailows electron flow to reoccur, after being inhibited, it suggests that the inhibitor added at
point A blocks electron flow from complex I to complex 111 (recall, succinate wiil add electrons at complex II, blpassing complex 1). The only inhibitor in the list that does this is rotenone. Antimycin A blocks electron flow from complex III to complex IV Atractyloside blocks ATP/ADP exchange across the inner mitochondrial membrane and will stop electron flow due to an inhibition of phosphorylation. The addition of succinate ri,.ould not be able to overcome an inhibition of ATP s1-nthesis due to lack of substrate (ADP). Dinitrophenol is an uncoupler,

+l H"N_C-H "l
CHz
I

CHz
I

CHz

cooo-Ketoglutarate

cooGlutamate

Panel A indicates the general reactton for a transamtna[ton reaction whereas Panel B shows the [ransamlnation between aspartic acld and
crt,-ketoglutarate.

Tl\
- --. i.'.ruld

C'':.-: ''.:-i Cr-::"--''

:--,-:

97

'-. ...thi.h

:r:t1ce of

in the not a1low eleclron flow from complex-1 oxidizable subrotenone Lactate is another

18

would not overcome the block of ,electron

wr11 gener-

.- '-er from complex I as lactate oxldation I' . ,,rrH, which adds electrons to complex

rheanswerisE:lnhibitionoftheprotontranslocating ::-' ffi;;. oligomlcin blocks ihe F' componen: prtrit-: '' therebl- blockrng pt"i""-"^"*I.n,i'-'g ATPase' olig"mr ho* through the enz)'me and ATP s)nthesis
cindoesnotaffectanyothercomplexolorrdatrl-ephosphoryladon.

tn oxygen Tk answer is D: Dinitrophenol' The increase (which is allowrng electron stimulated by succinate to oxygen) ts berng blocked by oh,

-:-:i. - ';";.;;pl.*it AfP . ., ..ir-,, which inhibits ATP qmthesis The block in consumption .-:.tt; I.ud, to Lhe cessation of oxygen
-

t9 fhe

gluconeogenesrs from .nrUo"ytur. deficiency will lmpair

answer is A: Fasting blood glucose'

p'vrur-ate

i;;;ffi ;;;

the link - - . .rnthesis, is an uncoupler' whic!-uncouples production Diniand '- .'.3.n oxygen consump[on of answers Note ihe onlyuncoupler on rhe list :-enolis increased as ar, ,fr. ,u,a oi o"ygtoconsumption has or succlnate was when either NADH - ,::-:ared to that grais due to the lack of a proton -, .--.ir-tg .Uorons. Thls there is no "back . .l-'t"ifr. p.esence of an uncoupler' so the electron flow is :::ure" to oxJgen consumption' and I uncoupler' tl-,.rn in thJ absence of the

-'

,.1,i-t.

coupling of ondation and phosphorylation of :;;;hai .# u[o- electron now' in the absence

The

h'vpogivpyruvate, iht"bv leading to fasting dehydrogenase defi..rr'tru -or. easily than u py"t"tt generale wril primarily affect the abillty to

.i.".y t*}tr.f,

i;;;

;;,"t*,

-.

,t{

;;i ;;]^;y ;;;;;";t,

carboi-'vdrates)'qlanine amino rvhich i" the blood ls a measure of liver damage' not distlnguish between the two possrbihtres' would be the same under both

transferase

acid 1eie1s during fastlng conditions'

1eve1s'

as

would lnsulin

and glucagon

--..

Lvi 20 The answer is B: Metformin blocks hepatic of hepatic leads to a reduction ,"r"tit. is accomplished- through the
"etformin This

gluconeo'

dehydrogenase' The Tte answer is : Cytoplasmic malate is required in liver as - - -:1;sm1c malate dehydrogenase shuttle in rransfernngreducrng - .* .'i r#*rfate/aspartate
.,.-i-alents across the

e for an inactlvatlng mutation - ..:ose. The same is tru iipyruvate kinase were defecwe' be converled to lactate v''ithout -: rmulale, which cannot first A defect Cly*t"l-l the glycerol-3-phosphate ...'.-.-arZg"r-ture will prevent electrons t the mitochondnf - -.:rrie from transterring

.'';;;.. (since the .l..not's cannot be lransferred ,:. ;;;;;; wll1 lead to the reduc- *r,i.tlorrdnal matrk) and ,:'"ipt""*,e ro lactate to regenerate *oo --l:l:,tY glucokinase will hlock -..;iJt;;^.t ""t A defect ii-rlactate formatlon from ..:.L;,';;h ,-,o p1o-tuutt or

ln inner mitochondrial membrane wil1bui1d up of such an acti\'lty' NADH Ieve1s.

!i.,.on"og.r-tesis. protein kinase' whlch activation of the AMP-activated within the cytop}asm phosphorylates and sequesters of CREB activiry (a TORC2, whrch is a coactivator two g1ufactor needed lor expresslon of

- .i-ir

t:t Yll

-,*;r*tate
for thls

li::*:::

trur-tr.riptio,-t and glucoseconeogenic enzymes' PEP carboxykinase is absent from the O-pfl"Ipfl.r,*tl ffl"t, rn'-hen TORC2 as the s)'nthesis of nucleus, gtu.or'"ogt"esis is lmpaired One of the malor two key enzymes L g"ut1y reduced lut'u"' generated from gluconeogenLc precirsorc i' muscle ln the Cori if-t. ,.a Uiood telt' and exercising fo one glucose' which cycle, two laclates are converted is blocked' lactate ,; d*" exported' If gluconeogenesis increase' and potenis not utllized and i"ts levels can However' in the absence of tia11y lead to lactic acidosis insuffici'ency' thls does congestive heart failure or renal of musThe heart, w'ith its masslve amount

-:--.'.rL\,buttheli.verusesprimanlythemalate/aspartate lor an overIJe activity See the figure below

-:-,r- of the maiate/aspartate shuttle system Cytosol Glucose

,ro,"o..rrr. utilize the lactate for energy c1e and mitochondria, can or has losl muscle unless the hearl is dysfunctional functional kidneys can also overcome

*urr'

Cooa,

Mitochondrion

f*, --Apyruvate
*oo*

fr-

uno'1'+Malate

-____> rvlarate

_r

NAD-

*'"T"1: ,o Aspartate

f,ro,u,urnu,"

flL_

Oxaloacetate+/

\-+ NADH'/

)o
ElectrontransPod chain

Glutamate

Aspadate

lrfier mitochondrial membrane Answer 17

98

Chapter t 1 the lactate lmbalance caused by metformin treatment. Metformin does decrease the lnsulin resistance, but this does not increase lactate in the aerobic state. Metformin
does not inhibit the TCA cyc1e, glycolysis, or dietary proteln absorption. These interactions are outlined in

the figure be1ow.

fLX-1-l

Ol FMPKiS@-eo++-6J
Metformin

ffil

.--.--+: :
l

tratraz-l
Po+3Sequester in
cytoplasm
:

I
, I

t_ l-tCREP

:
:

: i Glucoseexport
Enhanced

PGCI cr expressron V lncreased gluconeogenic gene expresslOn

I

t
I

{:'- -r |

gluconeogenesis

Nuclear membrane

TORC2 sequestration in the cytoplasm after phosphorylation by the AMPactivated protein klnase, which is activated by metformin treatment. This Ieads to reduced synthesis of key giuconeogenic enzyrnes, thereby reducing
gluconeogenesis in the liver.

Chapter 12

Glycogera Metab*eisrm
student cn vcrisu's :.?- qtizzes : :lic i1'tth *is';he degradatiaw of tht and staragt molccule -in tfu bafi'' , :lrrcliate :i tlrcse prac"ises is also hty as is iirr .,tng of tlie nutltitttde of disutsts thst -.'tt ritelalolisllt.
.
error s,ymptoms. At the pediatricians office, an inborn ol metabolism r,r,:rs consiclered, r'vhich could explain the hl.pogl,vcemia \\ihlch explanatron is n'rost 1lkei1?

(A) Frucrose rnhrbition ol the debranching enz,vme (B) Galactose -1-phosphate inhrbitron ol glycogen phosphorylase

(C) Fructose-1-phosphate inhibition ol gllcogen

phor,\'ln5s

pl-Los-

: TlONS
r

(D) Fn-rctose-6-phosphate inhibition ol glycogen phosphory'lase

I :rgle best answer.

due :-.n-o1d inthnt u'as brought to the pediatriciar.r lone . . - c u-eakness (m1'opathl') and poor muscle

(E)

Galacrose inhibrtlon ol aldolase

-.,.:'t. ancl heart farlure. Tl're rnfant had

liver -':lra) Ph1'sical exam revealed an enlarged

.-,rd lailure to thrive , and had breathrng problems l.racl

chilcl cran clue to fussiness ancl a tender abdomen The

seemecl

6-month-old lnlanr u'as broughl to

c1o r.l'e11

to rhe pediatn-

a1w'a,vs l'ed

increasecl

to

Lhe time benl'een teedrng r'vas more than 3h. Ihe baby alu'ays seemed

until
i.f

trouble holcling up hrs head Blood lr'ork -,',.i earlv Liver failure. A 1lr'er bropsy indicated that :::1 \\'as present and of normal struclure A poten, : .CL 11-t thls child is rvhich of the fo110r'l'ing?

hungn' ancl irrltable

not fed lrequently Upon exani-

notecl' ,-t.tio,-r, hep;rtome gal)' and enlarged kidneys rvere Subseand blooci r'vork sholvecl lasting hlpog\'cemia laboratorl'analysis demonslraled that rn response quent

' .r gll'cogen PhosPhorl'lase

'.-er gl,vco gen sYnthase

tl n gl.,.ugor-l. cirallenge,

only about 10% of the normal oig1,,,core u'as released into circulatron' lr'hich ^-o.,r-r, signihcantlf contributed to the fasting hypoglycemia'
hkely? fl,it-ri.t-, enz).me defect in the patient is the most (A) Glycogen sYnthase

',

er o,-glucosrdase

-lr-er

--' er clebranching enz)me branching enz)me

--

to the pediatrician due to 1-'ut :1.- exercise intolerance. In gl,rr class, the bo1'
ear-o1d boy is brought
.

-: ol Lhe folloll'rng :

or'ved rie u,ith anaerobtc actlvitres Laboratory tesls sh ,,,li of lacLate procluction uncler such conditions The r'vhich \\'as e\renturlly forrr-,d lo hal'e a mutalion in
enz1r11s52

(B) Branching enzyme (C) Debranchlng enz,vme (D) Glucose-6-PhosPhatase (E) Fructose-1,6-bisPhosPhatase
[ar1A 'l-nlonrh-oid inlant is seen b,v the pediatrician for shor'vs distinct hepatospleure lo thrive. Examinatlon and nomegalli Lab resuks shor'v elevated transaminases ol liver failr-rre The boy dies shortllbrllrubln, suggestive carb oh,vdraLe there alter, rr-t.1 .,p.rr-, autop sy, precipiute d the liver' The boy most likelr-had r,r-as found throulhour a mutatlon in rvhich of the follolving enz,vmes?

Lir-er glYcogen PhosPhorYlase Lr.ver PFK-1 \Iusc1e PFK-l \tuscie glucose-6-PhosPhatase Lir,er glucose-6-phosPhatase

diet -- l-month-o1cl infant, r'vhen swi'tched to a formula juices, begins Lo vomit and displays severe :.is lrult fruit .lulces :.. pogi,vcemia after earing. Removal- ol the

-].,r-r'ii.. diet seemecl lo reduce the severiLy of

the

(A) Giycogen PhosPhorYlase (B) Debranching enzYme (C) Glycogen slnthase (D) B-glucosrdase (E) Branching enzYme

99

100

Chapter 12

.,..$.. An inactivating mutation in which of the following proteins can lead to fastlng hypoglycemla? (A) Liver PFK-I (B) Liver glucokinase (C) Adenylate cyclase

'il

Patients with von Gierke disease display hepatomegalr Glycogen content in the liver is increased, relative tc normal, due to which of the followlng effects of glucose6-phosphate in these patients?

(D) Galactokinase (E) Fructokinase

,.:1.,:,,

If the turnover number ol all enzlrnes involved in g1ycogen metabolic regulation and activity is 100 reactions per second, how many glucose molecules could be removed from glycogen in I s upon actiyation of one molecule of protein klnase A (PKA)?

(A) Inhibrtion of phosphorylase a (B) Stimulation of phosphorylase b (C) Inhibition of glycogen s).nrhase I (D) Stlmulation of glycogen s),rrrhase D (E) lnhlbition of glycogen phosphorylase kinase

IZ

The hlperuricemia observed in patients with von Gierke disease comes abour due to which of the following?

(A) i00 (B) 1.000

(A) Glucose-6-phosphate inhibirion of kidney tubule

absorption o[ urare

(c)

10,000

(B) Lactate rnhibition of kidney tubule absorption of

urate

(D) 100,000 (E) 1,000,000

:1$.:, An individual is taking a serene walk in the park when he spots an escaped alligator from the zoo. The indlvidua1 runs away as fast as he can. Glycogen degradation is occurring to supply glycolysis with a substrate even before epinephrine has reached the muscle. This is due

of glucose-6phosphate dehydrogenase acriv,try (D) Glucose-6-phosphate stimularion of glycogen synthase D

(C) Giucose-6-phosphate inhlbition

(E) Glucose-6-phosphate acrlvarion of amidophosphoribosyl t ransferase acrrviry

to which of the following?

l3

(A) Sudden decrease 1n blood glucose 1eve1s (B) Increase in sarcoplasmic calcium levels (C) lnsulin binding ro muscie ceIl receptors (D) Decline in ATP levels (E) Lacrate producrion

Consider the case of an athlete who has just completed work out. At this point, the athlete consumes a sports drink, which contains a large amount of glucose, which enters the circulation. Glycogen degradation ls inhibited in the hver under these conditions, prior to insulin release, due to allosterlc inhibition of which of the folIowing enz).rnes?
a

.,.$.

''"''

As the lndividual in the previous question continues ro *., from the alligator, the muscle begins to imporr g1ucose from the circulation. This occurs due to which of

the following?

(A) Insulin binding to muscle cel1s (B) Epinephrine binding to muscle cells (C) Glucagon binding to muscle cells (D) lncrease in intracellular AMP ier.els (E) Increase in intracellular calcrum levels
.LS.: An 1S-year-oid man visits the doctor due to exercise intolerance. His muscles become stlff or weak during exercise, and he sometimes cramps up. At times, his
urine appears reddish-brown after exercise. An ischemic forearm exercise test indicates very 1ow lactate production. A potential enzyme defect in this man is which of the following?

(A) Glycogen s),rthase I (B) Phosphorylase kinase a (C) Phosphorylase a (D) Protein phosphatase 1 (E) Adenylate kinase

l,*

A muscle cell line has been developed with a nonfunctional adenylate cyclase gene. Giycogen degradatron can be induced in this cell iine via which of the following
mechanisms?

(A) (B) (C) (D)

Addrtion of glucagon Addition of epinephrine

lncrease in intracellular magnesium lncrease in intracellular AMP
Increase in intracellular ADP

(E)
.1.$,

(A) Muscle glycogen phosphorylase (B) Liver glycogen phosphorylase (C) Lil,er PFK-I (D) Muscle glucose-6-phosphatase (E) Muscle GLUT4 transporters

researcher created

a liver

ce1l

line that displayed

was

very low levels of glycogen. The glycogen rhar

rynthesized was of normal structure, but the overall levels of glycogen were about 5olo of normal. Which of the

foliowing is a potential alteration in the cel1 line that

would lead to these resuits?

GlycogenMetabolism (A) An altered glycogen

UDPglucose
s)-nthase with a reduced

l0f

for

16

(B) An altered phosphorylase kinase with an increased

for glycogen

(C) An altered UTPglucose-1-phosphate uridyl transferase with a decreased K- for glucose-1-phosphate (D) An altered glycogenin wlth an increased K- for
UDPgiucose

Ten hours into a fast, in a normal individual, u-hrch I the following best represents the acti\.1ty and phosphr.rylatlon state of a number of key enzymes wrthin rhe liver?

(E) An altered phosphorylase kinase with an increased

K_ for glycogen s).nthase

PFK.1

Glycogen Synthase

Phosphorylase Kinase Phosphorylated?

Yes

Pyruvate Dehydrogenase

Active?

Phosphorylated?
Yes

Active? Phosphorylated? Active?

Active? Phosphorylated?

iA) No
,B)

No

Yes
No
Yes

Yes
Yes Yes Yes

No

Yes

Yes

D) E) ll

No No No No
Yes

No
Yes

No

A woman with nonclassical galactosemia is considering becoming pregnant and is concerned that she wlll be unable to synthesize lactose in order to breast-feed her child. Her physician, who recalls her biochemistry, te1ls her this should nor be a problem, and that she ,,vi11 be able to synthesize lacrose at rhe appropriate trme. This is true due to the presence of which of the
foliowing?

used to produce glycogen in the 1iver. Which one of the foliowing liver enzymes is required for this conversion to occur? (A) o-ketoglutarate dehydrogenase

(A) Galactose-1-phosphate uridyl (B) Phosphoglucomutase (C) Fructokinase

(D) Aldolase

transferase
aor

(B) Pyruvate carboxlrlase (C) Pyruvate kinase (D) PFK-] (E) Glucose-6-phosphatase
Your patient is a marathon rllnner and has vislted your office to ask you about carbohydrate loading to increase his performance during a race. For a fu11 week prior to a race, he eats three meals a day of pancakes, potatoes, brown rlce, and pasta and does not exercise at a1i. He has not noticed any success with this reglmen. Which of the following answers best explains why he is getting no benefit from his "carb loading"? (A) Carbohydrate loading is a myth

(E) Phosphohexose

isomerase

The energy required to store one molecule of glucose6-phosphate as a portion of glycogen is which of the following?

(A) One high-energy bond (B) Two high-energy bonds (C) Three high-energy bonds (D) Four high-energy bonds (E) No high-energy bonds

(B) He

is not

depleting glycogen srores prior

ro

loading

(C) He is not on the carbohydrate loading diet long

enough prior to the race

(D) He ls eating the incorrecr foods for carbohydrate

An individual has been eating a large number of

oranges durlng the winter months to protect againsl getting a co1d. The excess carbons of citrate can be

loading

(E) He is too highly trained as an arhlere for anythlng to

increase his performance

ANSWERS
fhe answer is C: Liver u'glucosidase' The r'nfant has Pompe drsease, a loss of liver u-glucosidase actir-it1r of norThis is glycogen slorage disease 1l The hnding mal gl1'cogen struc[ure ehminates lir''er debranching

enzyme

lvith gly.og.r-t accumulates in the lysosome., interlering

is a lysosomal enzyme, and

nondegraded

The mrsstng and b".ar-tching activities as berng deficlent

iyroto*ut function (hence, a lysosomal storage disease)' ihe malfunctroning ol the lysosomes is r'vhat leads to the phosphomuscle and liver ptobl.,rt. A defect in glycogen and an rylase (liver) .toul,l 1"ad to fasting hypoglycemia.' *tu.g..l liver, but not rhe muscle problems exhibited by

Glucose

Site of enzymatic action Lvsosomal g[ucosidase

Pompe disease (heart)

nfi
I

u,To\

liloy-6,toto
l-

1.

sr*ffo

McArdle disease (mu

Phosphorylase

Transferase

Cori disease (liver, muscle)

)oo(

Debrancher

crPSG1orer-ofl f11{-crtofgf c.kofo

ffi McArdle disease (muscle) xlx
Phosphorylase Glucose-1-POo Glucose-6-POo

)Ofi
I

Glucose-6-Phosphatase

I
Glucose
diseascs' G\'togen phosphothat are deficient in various glycogen storage glycogen and an indrcation of some of the enzymes of glucose The debranching enzyme f ransfers a sma11 number in glycoger-r, ,.t.rrir.g glucose-1-phosphare. r1,1ase hydroiyzes rhe cr-i,+linkages remo'es the cr-1'6-linked enzyme also to a longer i-rlir. of sugars (reaction 1). The d.ebranchrng residues from branch pornrs and adds them
o1

The carabolism

sugaralrheoriginalbranchpoint(reaction2) O:.,.glucose-1-phosphateisconrerredioglucose-6-phosphate,glucoseisreleasedbythcactron withln lysosomes by acid ct-glucosidase

of glucose-6-phosphatase

,i

r-rli

proporlion of g1y.og.t is toially degraded

Glycogen
s)'nthase lvould also lead --,-',rnq hlpoglycemra, but would not lead to severe *-.- : and liver disease. Additionally, rn an indir.idual
-

Metabolism f 03

: - 'ld. A defect in glycogen

- -'. lefect in gll.cogsn

r,vhich is exported (10% of the expected) is derived from

s)'nthase, glycogen would not

. . '.rd in the liver biopsl. since it could not be formed. --. -=ure on page 102 summarizes steps involved rn - , -i.n degradation, and the glycogen storage disease -- ::sults if an enzyme is defective.
-

the actn.ity o[ debranching enzyme, r,vhich hydrolyzes an o-1,6-glucose hnkage, r.vhich produces free glucose. The hepatomegaly arises due to excess glycogen

in the

1ir.er (glucose-6-phosphate

will

actir.,ate glycogen

s),nthase D), as does the increase in kidney size. A plcture ol a 25-month-old untreated child rvith this disorder ls showrL below'. A lack ol glr-cogen synthase $.ould

Tle answer is

C: Muscle

PFI(-1. The child has a form of

. --ien storage disease known as type VIi, Tarui dis,,:: .,r'hich is a lack of muscle phosphofructokinase 1 : :.-ll acti\.ity. The lack of muscle PFK-I means that . - --,i srs rs imparred, so anaerobic acti\.ities are signifi.in such rndrvrduals. Slow aerobic actir-i:: -.rhich can be powered by lattv acid oxidation, are - -::-:1 in such children. Strenuous aclivil)' will lead ::-.scle damage and weakness due to thls block in . --. rsis. Glucose-6-phosphatase is only found in the .: .and to a small extent, the kidneli, and a lack of :-- . activlty r,vould lead to fasting hypoglycemra, but .,.-C not affect muscle glycolytic activitl A delect in .: PFK-1 acti\.ity would not affect muscle glycolysrs. -- *riect in liver glycogen phosphorylase rvould also r..: io lasting hypoglycemia, but r.vould not alter the .,.. ..i muscle giycolysis, or lactale formation from that
I : -1\\-3\'.

not lead to hepatomegaly, ri-hi1e a lack of branchrng enzyme ieads to a different g11 cogen stora5le disease, with very dlfferent s)rrnptoms. A lack oi debranching activity would not lead to hepatomegalr' :.inc1 r,r,ou1d
allor'v more glucose release than is obsen-ed through ihe

normal action of glycogen phosphor,vlase A delecL in

fructose-1,6-bisphosphatase would lmpair gluconeogenesis, but should not affect the abrlrty of gly-cogen to be degraded to raise blood glucose levels.

, . , . curtailed

The answer is (: Fructose-l-phosphate inhibition of glycogen phosphorylase. The child has hereditary r---iose intolerance, a defect in aldolase B actn'rty in -:-: liver. This leads to an accumulation ol fructose-1: -)sphate in the liver (and, as fructokinase has a high . . a large amount of fructose-l-phosphate accumu-

A 25-month-o1d child uith von Gierke clisease. Note the hepatomegaLy and eruptive xanthomas on the arms and legs. The child is in the third percentile for height and u,eight, indicating a failure to thrive.

,,.:s). At high

ler.e1s, fructose-1-phosphate, through

..:::ilarity in structure to glucose-l-phosphate, inhibits :.-. cogen phosphorl4ase acti\.ity, Ieading to hlpoglyce..--: (glycogen degradatlon is inhibited when blood g1u---se levels drop). The fructose is denved lrom the fiuit -.rces introduced to the childs diet. Fructose does not
:--..s

The answer is E: Branching enzyme. The child has a lack of branching enz;rme actMty, another glycogen storage disease, tlpe IV (Andersen disease). In this case, the
glycogen produced is a 1ong, straight chain amylopectln,

which has limited solubility, and precipltates ln the liver

(recall, the lir.,er has the highest concentratron of glycogen of al1 tissues). Thls leads to early liver failure (thus, the high bilirubin and transaminases in the serum) and death if a 1ir.er transplant is not performed. Defects in any of the other en4rrnes listed would lead to a different climcal scenario. Lack ofglycogen phosphory,lase or slrlthase. within the 1iver, would lead to fasting hlpoglycemia, but not liver farlure. Lack of these enzyrnes in the muscie would lead to exercise intolerance but would not aflect blood glucose levels. Lack of o-glucosidase is Pompe disease, which also leads to an early death, but is due to the lack of a lysosomal enzyrne, and there is no giycogen precipitation wrthin the body of the liver. A lack of debranching activity is glycogen storage disease 1II, but w.ould also lead to fasting hlpoglycemia, without glycogen preclprt3tion within the liver. A number of the glycogen storage diseases are summarized in the figure on page 104.

::hrbrt debranching enzyme, and fructose-6-phosphate no effect on glycogen phosphorylase (reca1l, one of ::e products of the glycogen phosphorylase reaclion rs

.-'-icose-1-phosphate, not glucose-6-phosphate). Galac--.se is found in iactose, which, whiie present in mi1k, is

::rt found in fruit juice.

The answer is D: Glucose-6-phosphatase. The child ::as Von Gierke disease, glycogen storage drsease type ,. a lack of glucose-6-phosphatase. In such a disorder, .Lucose-6-phosphate, whether produced from glycogen :egradation or gluconeogenesis, cannot be dephosphorlated lor glucose export, and the liver cannot main.iin blood glucose 1evels. The small amount of glucose

LO4

Chapter 12

Glycogenosis
Hepatorenal 9., Gierke disease

Deficient
glucose-6-phosphatase

Biochemical diagnosb
Normal glycogen; excessive amounts in liver and kidneys Normal glycogen, excessive in all 0rgans

Glinical symptonn
Hypoglycemia, hyperlipemia, ketosis, hyperuricemia, hepatomegaly, dwarf ism Muscle hypotonia, heaft failure, neurologic symptoms, infant death Hepatomegaly, hypoglycemia; mild course of disease

Generalized, malignant g.; Pompe disease;

cr-1 ,4-glucosidase

cardiomegalia glycogenica Hepatomuscular, benign g.;

Cori disease, Forbes disease

Amylo-1,6-glucosidase

(with subvariants 3b through f) Liver, cirrhotic, reticuloendothelial g.; Anderson disease: amylopectinosis
cr- 1 ,4-glucan:

Abnormal glycogen, with short outer chains, in liver and (more rarely) in muscles Abnormal glycogen, with long outer chains, in liver, spleen, and lymph nodes Normal glycogen, excessive amounts in muscle
I

ct-1,4-glucan6-glycosyltransf erase cr-glucanphosphorylase of the muscle ct-glucanphosphorylase of the liver Phosphofructokinase

of the muscle

Cirrhosis of the liver;

hepatosplenomegaly

l\iluscular 9., Mcardle-Schmid-Pearson disease Hepatic 9., Hers disease

Generalized myasthenia and

myalgia, myoglobinuria Hepatomegaly, relatively benign

Normal glycogen, excessive amounts in liver Normal glycogen, in the skeletal muscle Normal glycogen, in the liver

Muscular g.; Tarui disease

Muscle cramping, myoglobinuria Clinically mild manifestation, hepatomegaly, hypoglycemia

Hepatic g.; X-chromosome inheritance

Phosphorylase-b kinase
of the liver

Answer 5: A summary of the glycogen storage diseases

nor increase, and pK{ will stay inactive) Under such conditions, only the allosteric effectors in llver will be actlye, and there is no activator of glycogen phosphorylase b. When the hypoglycemia rs severe enough, epinephrine release, working through its s-receptors, will acrivare phosphoiipase C, leading to calcium release. The increased calcium can activate phosphorylase kinase, which will activare phosphory1ase, but fasting hypoglycemia wiil still occur. Defecrs in Iiver PFK-1 or glucokinase will not affect glycogenolysrs or gluconeogenesis. Defects in liver galactokinase or fi-uctokinase will not a1low for metabolism of gaiactose or fructose, but do not affect the ability of the liver to degrade glycogen, or perform gluconeogenesis from other precursors. (cAMP levels
wr11

The answer is (: Adenylate cyclase. Ifadenylate cyclase is defective, glucagon cannot initiate the activation of glycogenolysis and inhlbition of glycolysls in the hver

phosphoryiase molecule can release 100 glucose residues

per second from glycogen, and since there are 10.000

actrve phosphorylase molecules, 1,000,000 molecules of glucose are released per second once a srngle molecule of PKA has been acrivated. This is an example of cascade

amplification, in which an increase in activity of just one molecule at the top of the cascade can result in a large response further down the cascade.

The answer

is B: lncrease in

sarcoplasmic calcium

levels. When the individual begins ro run away from

the alligator, muscle contraction leads to calcium release from the sarcoplasmic reticulum to the sarcoplasm. This increase in sarcoplasmic calcium binds to the calmodulin subunit of phosphorylase krnase and activates the enz)-rne

in an allosteric manner, in the absence of any covalent modification. The activated phosphorylase krnase will phosphorylate and acrivate glycogen phosphorylase, which will lnitiate glycogen degradation. When
epi_

The answer is E: 1,000,000. One acrlve pK{ can activate in 1s 100 molecules of phosphorylase kinase. Each phosphorylase kinase can, in 1s acrlvare 100 molecules

of glycogen phosphorylase (so at this point we have 100 tlmes 100 active molecules of phosphorylase, or
10,000 actlve phosphorylase moiecuies). Each active

nephrine reaches the muscle, phosphoryiase kinase will be fully acrivared ua phosphorylation by pK{. The activation of glycogen degradation under these conditions is not due to a decrease in blood glucose levels, insu_

lin binding (insulin would not be

released under rhese

conditlons), a decline in ATp levels (the AMp-activated

GlycogenNletabolism 105

Extraceliular

Cytoplasm Muscle contraction

otrflt||Iry

,::

::- r:,. ' .,-i lo calcrum release '' , :ir:-'rlasmic reticulum,

"

a . .::iation of glycogen . : ::gradatronbycal.: :-iscle. Muscle conSarcoplasmic reticulum

--rva ^^2+

O f C"fr"Orf * \ .y Grycogen .Yn,nu.i 1 {inactive) dependent \-----t( -p (.!!lgi_Ir9:U '\,: ...-''> / synrhase
...\o,y"og"n (active)
/\tphosphory,asea-p -, ) phosphoryrase \._ _ _,/" " ""i;.;;";" " urycogen \ phosphorylase
(inactive) Glycogen

Ca2*-calmoduIin
---

; * ;- :::-Jfiasekinase,and -.. -nhibrtron of glyco.1,.- - - - .-.-:.: and rhe aclivarion

:.:::gradation.

kinase does not activate glycogen degradation), .-:iate production, the end product of anaerobic ..-:.lo1ism. The figure above shows the stimulatton .-i-cogen degradatlon, working through calcium . -'--.:tion of the calmodulin subunit of phosphorylase

,:. -.-n

._*_:se.

TLe answer is D: lncrease in ---,1? ler,els increase

intracellular AMP levels. As in the muscle due to the need for

-.

1 :or muscle

-. ..::ein kinase is to increase the number of GLUT4

-r::1sporters in the muscle membrane, in a process simi.::o the action of insulin. This enables muscle to take -: glucose efflciently from the circulation when inter,-"- energy leveis are 1ow. The ability of the muscle to :,..a up glucose under these conditions is not due to an i-rrease in epinephrine levels, an increase in sarcoplas-

contraction, and the activity of the adeklnase reaction, the AMP-activated proteln kinase -,:e ,rrned on. One of the effects of the AMP-activated

shown in the figure below, there are many glycogen particles present in the muscle cel1s.;ust below the sarcolemma, as the glycogen is not able to be degraded. Muscle damage also results from vigorous exercise, releasing myoglobin lnto the circuiation, which is what leads to the reddish-brown urine after exercise. Alterations in liver enzymes (phosphorylase or PFK-1) would not affect exercise tolerance in the muscle. Muscle does not contain glucose-6-phosphatase, and this problem is not due to a lack of muscle GLUT4 transporters, as the muscle cannot utilize stored, internal g1u.or.
supplies.

CN

::--: calcium levels, or insulin binding to muscle cells. -:der conditions as described in the question, insu-:: uill not be present in the circulation to bind to the :-uscle ce11s. As the muscle does not contain glucagon
iceptors, there is no effect on muscie when glucagon is ::esent in the circuiation.

The answer is A: Muscle glycogen phosphorylase. The

:atient is lacklng muscle glycogen phosphorylase .rd cannot utilize muscle glycogen for energy This -s another glycogen storage dlsease, type Y McArdle
-...'h) lactate

jrsease. The lack of muscle glycogen phosphorylase is production during exercise is very 1ow. As

The electron micrograph demonstrates an abnormal mass ol glycogen (not surrounded by a membrane) particles just beneath the sarcolemma, which distlnguishes this disorder from Pompe disease (a iysosomal disorder in whlch glycogen wlthin the lysosomes cannot
be degraded).

f
I

06 I

Chapter 12
The answer is D: Stimulation of glycogen synthase

D. G1y-

cogen slnthase D (the rnactive, phosphorylared form) can be allostericaily activated by glucose-6-phosphate blnding to the en4rrne. Glucose-6-phosphate will inhibir rhe AMP-stimulation ol muscle phosphorylase b, bur does not have any allosteric elfect on the other enzynes llsted (PFK-1, glucose-6-phosphatase, or GLUT4 rransporrers) as answer cholces for this problem.

acti\rate gl).cogen phosphorylase b; the allosteric actlvator is specific for AMP The table below summarizes the ailosteric interactions rnvolved in glycogen metabolism.
Form of

enzyme

Phosphorylase a

Tissue Liver Muscle iver Muscle

I

Activator
Already active Already active None

lnhibitor
Glucose Creatrnephosphate None ATP. G6P

l2

is B: lactate inhibition of kidney tubute absorption of urate. Patients r,vith von Gierke disease
The answer
display eler,,ated levels ol lactate, which interferes u.ith the kidnev's ability to remove uric acid lrom the blood and place it in the urine This leads to h;rperuricemia The reason lactate levels are elevated is that the high glucose6-phosphate in the cel1 (reca11, the defect in this disorder is a lack ol glucose-6-phosphatase acriviry) forces glycolysis foru'ard, producing pyruvate, which is converted to lactate ln order to regenerate NAD- to al1ow glycolysis ro continue. Glucose-6-phosphate does not inhibit glucose6-phosphate dehydrogenase (that enzyme is regulated by the NADP. levels), nor does it regulate a commiued step of de novo purine s),.rlthesis, amldophosphoribosyl transferase (which ls regulated by adenine and guanine nucleotides) Glucose-6-phosphate does stimulate glycogen s)Trthase D, but that actitation does not play a role in eler.,ated urate levels. Glucose-6-phosphate does not affect urale absorption lvithin the ktdney

Phosphorytase b

AMP
Ca2,

Phosphorylase
kinase

Liver Muscle Liver

and

Glycogen

:!'.*""
|

and Glucose-6Muscle phosphate

None

The answer

increased ff for UDPglucose. A recluction in or.erall glycogen synihesls suggesrs that the biosynthetrc pathway is defectn-e in some srep. AIl glycogen

is D: An altered glycogenin with

an

molecules have, at their core, a glycogenin protein

ol six glucose residues, using UDPglucose as the carbohymolecule, which autocatalyzes the addition
drate donor. This strucrure then provides the initial pnmer required bi. glycogen synthase. 1l the K," for
UDPglucose is increased, the rate of lormation of glycogen primers will be decreased, as the levels of UDPglucose may not be sufflcient to a1low glycogenin to self-prime This r,vould result tn an overall reduction of glycogen levels withrn the cell. 1l a glycogen synthase had a reduced K,. for UDPglucose, then the enzyme would be actir..e at lower UDPglucose levels, and one lvould expect greater than normal glycogen synthesls. Phosphorylase kinase has as its substrate phosphorylase, not glycogen, so ans\ver B is not correct. If the uridyl transferase had a reduced Kn for a substrate, it would proceed at 1ow substrate levels and would not give the resultant phenotype. And, 1l phosphorylase kinase had an increased K,, lor glycogen synthase, then glycogen synthase would not be inactn ated as rapidly, and glycogen synthesis w.ould be expecred Lo continue under conditions where it should not. ieadlng to enhanced glycogen synthesis.

t'3

The answer is (: Phosphorylase a. The glucose in rhe sports drink will blnd to liver glycogen phosphorylase a and inhibrt lts acrivity allosterlcallyr Once the insulin signa1 reaches the hver, phosphorylase a u,-111 be converted to the dephosphorylated phosphorylase b by activated phosphatases. There is no allosteric rnhibitor for glycogen s)mthase 1, or protein phosphatase I (which is regu-

lated by protern inhibitor 1). Adenylate kinase is nor regulated allostericaliy, and there ls no allosrerlc inhibitor of phosphorylase kinase a (the nonphosphorylated
form can be activated by calcium).

14

The answer is D: lncrease in intracellutar AMP. AMP will activate muscle glycogen phosphorylase b allosterically, allowrng glycogen degradation to begin before any hormonal signal has reached the muscle. The addition of epinephrine to the muscle requires activation of adenylate cyclase to inittate glycogen degradatron, and adenylate cyclase has been inactivated in this cell line Muscle lacks glucagon receptors, so cannot respond to this hormone. An increase in intracellular calcium would lead to glycogen degradation (r.ia activation of phosphorylase krnase b), but magnesium does not have the same effect as calcium. Increases tn ADP levels will not

16

E. Under fasting conditions, rhe liver is exporting glucose, so the pathways of glycogenoiysis and gluconeogenesis will be active, rvhile glycolysis will be lnhibited (a11 due to the eflects of glucagon and activation of PK{). In glycolysis, PFK-2 is phosphorylated, activating rts phosphatase activir),, which leads to a reduction in fructose-2,6-bisphosphate ievels. This
fhe answer is

Glr,cogen

N,Ie

tlibrrlisl: l0
S:'.-".-

'..-. is ln a reduction of PFK-I activity (thus, PFK-I

:.iiir-e, but is not phosphorylated). C1;.cogsn dsg13-., ,-.n 11as been activated, and slnthesis rnhrblte d, via : :hosphorl.lalton ol glycogen s)'nthase, lnaetlr-al .. ,he enzyme (thus, glycogen synthase is not aetn-c, - , .s phosphorylated). P1-Losphorl'1ase kinase has been ' .'.rted, and phosphor,vlated, by PKA (so phospho- ,,se kinase is actne, and phosphorylated). Pyruvate
.-.:-, drogenase is rnactive under these conditions (clue -..ttr-acid oxidation in the mitochondria acetl-1-CoA . .i-. and NADH leve1s are high, u'hich slorvs dor'vn the -- c;-cle and inhibits p)'ru\.ale dehl,drogenase), and . aLso phosphor,vlated b1. tbe PDH-kinase, r'vhich is

is

und1.l transle rase. Patienis cannot me taboli-e

--

and the accumulating galactose-l-phosphate iniu: - .. w'ith gllcogen clegradatron. Nonclassical galact.,se::-. -, (t1'pe 2) is a deficit rn gaiactokinase, such thar g.rlac;.r.. cannol be phosphorl.lated. The compLications rn Lr:'. 1 galactosemia due Lo Lhe accumulaLion oI galactose 1 phosphate are not seen in ty'pe 2 galactosemia in eithtr case, lhe missing enz)'mes are not required for the srrr thesis oi lacLose See the {rgure below 1or both the pathr,va1' ol lactose s)'nthesis, and the delects in classical and
nr

rncl.ls5ir'al gall, l,,rcnt ta.

"7

..

ated

bi NADH

t8

The answer is B:

Phosphoglucomutase. For Lhis \\'oman , she needs io synthesize the pre:...rs UDPgalactose and glucose, both ol n hich '. "rrailable liom glucose . Glucose is conr.'erted tcr - ,.-'.rse-6-phosphate b1' hexokrnase rn the breast, r-,. then phosphoglucomutase u.i11 conr''erl this lo . -... se-1-phosphate (GlP) The GlP r,vill react u.rth ,,- in the glucose-1-phosphaLe uridr,l trnslerase r-..iion, producrng UDPglucose. The C4 epimerase .- then produce UDPgalactose lrom UDPglucose . The lPgalactose tl-ren condenses rvith lree glucose (usitrg . -,.rse sr,rrthase) to produce laclose and UDP The othe r ri-ifileS listed as answers are noi required to procluce --.r)se from the srngle precllrsor glucose. Fructoktnase : rnlqlle for fructose metabolism. Aldolase is a gl1-co- . enz)Tne, r.vhich is deficient in hereditarr. fruclose .-.-..lerance. Phosphohexrrsc isomerase ro\-crls glucose-''lhosphate to lructose-6-phosphate, r'vhich is not ::.lired lor lactose synthesis. Classlcal galacloserni.r

.,.rthesize

lactose

bond. For a molecule of glucose-6-phosphate (G6P) to be incorporated into g11.c6gen. the lollorving patl-nva). must be utrlizecl: G6P is convertecl to g1r-rcose-1-phosphate (GlP) vLa phosphogluc.rnlltase . the GlP reacts r,vith UTP to lonn
The answer is A: One high-energy UDPglucose via glucose- 1 -phosphate urid1,1 trans lerase. releasing p1,ropl-rosphate The resultant pl,rophosphate is h1'drol1.zed to two illorgar-ric pl-rosp1-rates, r,r,ith the loss ol one high-energ,v bond. The UDPglucose then reacts r,i-ith gi1'cogen to produce :r glycogen chain u,ith one additional sugar, and UDP is released. The overall ec3-ration lor these steps is. G6P + UTP + glycogen, yields

UDP

+ 2Pi +

(gl,vcogen),,*,

. These steps are outllned

belolv: Gluose-6-phosphate -+ Glucose- I -phosphate Glucose-1-phosphate + UTP -+ UDPglucose + PPi PPi + H,O -+ 2 Pi UDPglucose + glycogen. -+ C11cogen,,*r + UDP UDP + ATP -+ UTP + ADP Sum: Glucose-6-phosphate + ATP

:.iere. rype 1) is a delicit ol

galactose-1-phosphate

+ glycogen,, + H-O

glycogen,,*, + ADP + 2Pi

Glucose-'1-P

Galactose

l,-urP r
Y

l--arp V
Galactose- 1-P

Nonclassical galactosemia Classical galactosemla

oalactokinase

l_r rr,

- l.-r" nop

UDPGlucose
eoimerase ll

tl
UDP

,rr*t"Ji,,"3i",.*"

UDPGalactose ractose svnthase l r- o-Glucose (accePtor) o-lactalburin)

ll

Lactose
CH2OH

[ lt->UOp i

ePrmerase

Glucose 1----------.-//r IUDPI

\--_,

I .\(

galactose-1-P uridylyltransferase

Glucose-l-P + Glucose-6-P
|

Galactose

----

> Glycolysis
(other tissues)

1liue4

+
CH2OH

Ho,l-o.

q/" v
OH

Glucose

.-(o* )
OH

,l-o.on

Answer 17

Chapter 12
Glucose-6-PhosPhate

-----------+

ucose-

-phosphate

t
I

\
UDPGIucose

Fructose-6-PhosPhate

\
Fructose-1,6-bisPhosPhate

GlYcogen

Dihydroxyacetone-P

<----+

Glyceraldehyde-3-P

t
Glycerol -4GlYcerol-3-P

f I

GlyceratdenYde-3-PhosPhate dehydrogenase

1,3-bisphosPhoglycerate

t t
y', / /
Phos1hoenolPyruvate
phosPhoenolPYruvate carboxYkinase

Oxaloacetate

t \

Amino acids

Alanine

:rffiN-'r"l*"(
Lactate

Answer 19

1!'

fhe answer is A: cr-ketoglutarate dehydrogenase' ln

(whlch requires the partlcipation of u-ketog.lutarate dehydrogenase). From oxaloacetate, PEP carboxyki,-,ur. *i1i convert this to PEP, which will go through

citrare order for citrate to be converted to glycogen, the to oxaloacetate in the TCA cycle must first be converted

the gluconeogenic pathway up to glucose-6-phosphate' Froir there, GIP ls prod,uced, then UDPglucose' and glycogen' Pyrufina11y incorporation of the glucose into being a gluconeogenic enzyme' vate carboxyiase, while

in converts pyruvate to OAA, which is not requlred PFK-I and pyruvate kinase are this series of reactions. used in irreversible enzymes of glycolysis and are not

the gluconeogenic pathway

....ro"u", the pf,osphate from

Glucose-6-phosphatase

which is not required synthesized' See the figure when glycojen is being above lor the PathwaYs.
G6P,

other sources for energy to continue running ln the vernacular of the sport, when all the glycogen stores " is usuare exhausted, the runner "hits the wa1l This has shown a1ly somewhere around miie 20 Research that proper "carb loading" prior to a race can lncrease body stores of glycogen and increase performance' Though it is a sma11 increase (Iolo to 2ok), iL has been documented repeatedly in research studles even in hlghly trained athletes. Therefore, it is not a myth' g1yTo"properly carbohydrate ioad, one must deplete 2 to aog..t ,,o..t wlth very vigorous exercise about 3 iays prior to a race. This stimulates glycogen synnext thase whictr increases glycogen slores over lhe 2 to 3 days before it returns to baseline levels This is a critical step in the process of "overbujlding" g1ycogen stores. This is the step the patient is not dolng prJperly Vigorous exercise cannot then be continued

ar.i.g

the

zo

stores The answer is B: He is not depleting glycogen

glycogen stores

prior to loading. Marathon runners deplete their ,tor., of glycogen during arace andneed to catabolize

ilio*"

Pancakes, potatoes' rice, and pasta ate excellent sources of simple

i to 3 days of glycogen
will be utilized'

buildlng or the

carbohydrates.

Chapter 1 3

Fatty Acid Metabolism

.
"-.,

- ..abolism.

clnpter examines the students' ability to {.ate tlrcir hnowledge of fatty acidmetcrbolism .:ii clin.ical problems and carbohydrate

'"'.s

(A) Carnitine acyltransferase I (B) Carnltine acyltransferase II

(C) Acyl-CoA dehydrogenase

(D) Enoyl-CoA dehydrogenase

QL ESTIONS
lt:.ect the single best answer.

(E) B-keto thiolase

5'

I \ou prescribe ibuprofen to help reduce your patient's

-nflammation. Which of the following pathways is olocked as an anti-inflammatory mechanism of action
nonsteroidal antr-infl ammatory drugs? ,.-\) Prostaglandin slmthesis .B) Thromboxane slmthesis ,.C) Leukotriene slmthesis
,.1

A 3-month-oid child had her first ear infection and was feeding poorly due to the ear pain. One morning the parents found the child in a nonresponsive state and rushed her to the emergency department. A blood g1ucose level was 45mgldl, and upon receiving intravenous giucose the child became responsive. Further blood analysis displayed the absence of ketone bodies, normal levels of acyl-carnitine, and the presence of the following unusuai carboCic acids shown below The enz)rynatic defect in this child is mosr likely in which of
the following enz).rnes?

,.D)

A11

eicosanoid syrrthesis
release from the membrane

(E) Arachidonic acid

2 \bu have an asthmatic

o.
o).co' )p
O.

patlent who is already on an rnhaled steroid and albuterol, but is still having difflculqz You add montelukast to her regimen. Montelukast

cHr-cH2-cH2-

cH2

^/.o- c\\o

(Singulair) speclfically blocks the product of which of the following metabolic pathways?

- cnr-

cH2

vt vt vn2 vn2 - cH2 - cH2 -cH2 - cF -aao -^u t2-^u t2-^u -^u -n,O-

(A) Cyclooxygenase (B) Lipoxygenase

(c)

P450

(D) Cori cycle (E) TCA cycle

(A) (B) (C) (D) (E)

Fatty acyl-CoA s)Trthetase Carnitine translocase Carnitine acyltransferase I Carnitine acyltransferase II Medium chain acyl-CoA dehydrogenase

Coconut palm tress cannot survrve growing outdoors in Kansas. Which of the following is the best explanation for this finding?

Regarding the child described in questlon 5, why were fasting blood glucose levels so low?

(A) Coconut/pa1m oil is a saturated fat (B) Coconut/palm oil is a monounsaturated fat (C) Coconut/pa1m oii is a polpnsaturated fat (D) Kansas soil is not sandy enough to support growth (E) Kansas soil j.s too rocky to support growth

(A) Acyl-carnitine inhibition of gluconeogenesis

(B) Dicarboxyhc acid lnhibltion of gluconeogenesis (C) Insufficient energy for gluconeogenesis (D) Dicarboxylic acid inhibition of glycogen phosphorylase

(E) Reduction of red blood cell productlon of lactate for

gluconeogenesis

Anlnactivatingmutationinthe ETF:CoQ oxidoreductase will lead to an initial inhibition of which of the following enzlT nes in fatty acid oxidation?

A 6-month-old child presents to the physician in

hlpotonic state. The child has previously had
a

number

109

l0

Chapter 13
of hypoglycemic episodes, at which trmes blood glucose between 25 and 50 mg/dl Blood work shows
1er.e1s

12

1evels were

A mouse model has been generated as an in vit o system

ol ketone bodres (not elevated) during hypoglycemic episodes. Carnltine 1evels in the blood
normal
were, howeyer, elel'ated. Free fatty acid levels lvere also elevated. Liver and muscle biopsies show elevated levels of triglyceride. A 1ike1y en4Tnatlc defect is which ol the

lor stud;,ing fauy acid qmthesis An lnacrivating muta1ed to the cessation of fatty acid synthesis and death to the mice. This mutatlon is most 1ike1y in whlch of the following proteins?

tion was created which

followlng? (A) Carnrtine acyltransferase I (B) Carnrtine acyltransferase 11 (C) Medium chain acyl-CoA dehydrogenase

(A) Carnitine acyl translerase I (B) Carnitine acyl transferase 1l (C) Citrate translocase (D) Glucose-6-phosphate dehydrogenase (E) Medium chain acyl-CoA dehydrogenase
t3
u-oxidation u.ould be required lor the complete oxidatron of u,hich of the following fatty acrds?

(D) Hormone-sensrttr.e lipase

(E) Carnitine transporter

Carnitine deficiency can occur in a number of ways. Secondary carnitrne deflciency can be distinguished from prlmary carnltine def,ciency by measurlng which of the follorving in the blood?

CHs

cH3-

(CHz)n

-9|

t/icHr-cHr-Cr-

(A) Fatty acids (B) Acyl-carnrtine (C) Lactic acrd (D) Glucose (E) Ketone bodies

H-

,,-

B
generate

CH3-

(CHz)n

- CHr-?-Crr-Cro_
H-

?,.

,P

Which one of the followrng fatty acrds will the largest amount of AfP upon complete oxidation to carbon dioxide and water?

(A) (B) (C) (D) (E)

C16:0 cisA9 C16:1 cisA9 CiS:1 cisA6 C18:1 cisA9, A12 C18:2

cH3

(cHz)n

cH2

7n' ,p -cH2- ? -c.._

io-

CHs

l0

An indir',rdual contains an inactivating mutation in a particular muscle protein, which leads to werght loss
due to unregulated muscle fatty acid oxidation. Such an inactivated protein could be which of the following? (A) Malonyl-CoA decarboxylase (B) Carnitine acyl transferase I (C) Carnitrne acyl transferase 1I (D) Medium chain acyl-CoA dehydrogenase (E) Acetyi-CoA carboxylase 2
14

CHz IricHr-cHz-c. n (cHz)n cH3-Q- - - \..\_ | H" CHs CHs tt/i cH"-(cHz)"-c-cHr-C-c.

HH

l-l\o-

A 2-month-old infant wrth failure to thrive

tl

The net energy yreld obtained (moles of AfP per mole of substrate oxldized) when acetoacetate is utrlized by the nervous system as an alternative energ)'souree is which of the following? Consider that acetoacetate must be oxidized to four molecules of carbon dioxide during the reactlon sequence.

displays hepatomegaly, high levels of iron and copper in the b1ood, and vision problems This child has difficulty in carry,1ng out which of the followlng types ol reactions?

(A) (B)

17 18 1e

(c)
(E)

(A) (B) (C) (D) (E)

Oxidation of very long chain fatty acids Synthesis of unsaturaied fatty aclds Oxidation of acetyl-CoA Oxidation of glucose Slnthesis of triacylgycerol

(D) 20
21

15

A 55-year-old man had been advised by his physician to take Btmg of aspirin per day lo reduce the risk of

Fatq.Acid \letabtriisrn 111 blood clots leading to a heart attack. The rationale for this treatment is which of the following?

!8 An rndlvidual with a biotinidase

(A) (B) (C) (D) (E) t6

To reduce prostaglandin synthesis To reduce leukotriene syathesis To reduce thromboxane slnthesis To increase prostacyclin s)'nthesis To increase Llpoxin slmthesis

You are examining a patlent who exhibits fasting hypoglycemla and need to decrde between a carnitine deficiency and a carnitine acyltransferase 2 deficiency as the possible cause. You order a biood test to speciflcally examrne

rr, as sh.'.'i:l reduced rale \Ln :h. to produce fatty acids at a greatly absence of supplements) as compared to someone nh,r did not have the deficiency This is due to whrch ol the following? (A) Low actlvity of citrate lyase (B) Reduced activity of malic enzyme (C) Reduced activity of acetyl transacylase r D) DelecLive acy I carrier protein (E) Reduced abrlity to form malonyl-CoA

deficlencr-

the levels of which one of the following?

t9 Liver latty acid oxidation

leads to an enhancement of

(A) Glucose (B) Ketone bodres (C) lnsulin (D) Acyl-carnrrine (E) Carnitine t7
lnhrbitors specific for cyclooxygenase 2 (COX-2) were deemed more efficacious for cerlain conditions than
2A

gluconeogenesrs via u'hich of the fo1lor.vrng?

(A) Generatron ol precursors for glucose s1't-ithesis (B) Actj.r,ation of pyruvate carboxylase
(C) Actir.atron of phosphoenolpl'ruvate carboxl'kinase (D) Inhibrtion of pyruvate kinase rE) Inhrbilion ol PtK-2

35-year-o1d man

in

Ner.v York citl', originally lrom

lnhibitors which blocked both COX-I and COX-2 activities. This is due to which of the followlng? (A) lnhibiting COX-I increased the frequency of heart
attacks

Jamaica, purchased an illegally imported lruit from a street vendor and, within 4h of eating the fruit, began

vomiting severely When brought

(B) lnhiblting COX-2 did not alter prostaglandin production

(C) COX-2 is speciflcally i.nduced durlng inflammation (D) Specifically inhibiting COX-2 reduces the rate of
heart attacks

to the emergency department the man was severely dehydrated and exhibited several seizures. The toxic effects ol the fruit were interfering with which of the following? (A) Fatty acid release from the adipocyte
(B) Fatty acid entry into the liver ce1l (C) Fatty acid activatlon (D) Fatty acid transport into the mitochondria (E) Oxidatrve phosphorylatlon

(E) COX-I

ls ind.ucible and only expressed dunng wound repalr, whrle COX-2 ls expressed constitutively

'- i *

:n

O-

lI2
t

Chapter 13

ANSWERS
synthesis. Ercosanolds potent regulators of cellular function. They are derived from arachidonic acid and are metabollzed by three pathways: rhe cyclooxygenase parhway (prostaglandins and thromboxanes), lipoxygenase pathway (leukot_ rienes), and the cyrochrome p,150 pathway (epoxides)
are

lrom the membrane (whlch would block all elcosanoid

syrrthesis); however, they do interfere wrth the cycloory-_

The answer is A: Prostaglandin

genase

thromboxanes affect formation

pathwal Prostaglandins affect inflammation. of blood clots, and

(see the figure below). Nonsteroidai anti-inflammatory drugs (NSAIDs) do not block arachidonic acid release

leukotrienes affect bronchoconstdction and bronchodilatation. NSAIDs block prostaglandins as one of their anti-inflammarory mechanisms. Thus, while NSAIDS will block both prostaglandin and thromboxane syrthesis, it is the blockage of prostaglandin sprthesis which will block the inflammarory symptoms.

Arachidonic acid
Cyclooxygenase

Cytochrome P450

/
Prostaglandins

PGG2

HPETE

Epoxides

/
Thromboxanes

/\
HETE

Leukotrienes

Lipoxins

diHETE

/\

HETE

The answer

is B:

Lipoxygenase. Montelukasr

is

leukotriene blocker. Leukotrienes are formed through the lipoxygenase pathway and affect bronchoconsrriction and allergy pathways (see the figure in answer ro question 1). The cyclooxygenase pathway produces prostagiandins and thromboxanes. The p,i50 pathway produces epoxides. The Cori cycle rs related to gluconeogenesis (lactate transfer from the muscle to the liver), while the TCA cycle is utilized ro oxldize acetyl-CoA to CO, and HrO. The answer is A: Coconut/palm oil is a saturated fat. Saturated fats do not liquefy untll a much higher temperature than that at which monounsaturated or pol1unsaturated fats do (the melting temperature for saturated fats is greater than that for unsaturated fats). Conversely, saturated fats are solids at a higher temperature than unsaturated fats and cannot exist in a liquid form at a
lower temperature. Since the oi1 of a planr is irs "1ifeblood,,, at a lower temperature, a saturated oi1 would solidify and the plant would die. Saturated oil plants cannot sunrve in a temperate climate (Kansas) and need a tropical climate of warm temperatures all year round. Only pollunsaturated oi1 plants can sumve in a temperate climate (corn,

carbon-carbon double bond between carbons 2 and,3 of the fatty acyl-CoA, generaring an FADH2 in the process. The FADH2 then donates its electrons to the electron

A lack of the oxidoreducrase acri\1ry will lead to an

accumulatron of mttochondrial FADH2, depleting FAD levels, and reducing rhe activity of the acyl-CoA dehl drogenases. The lack of FAD does nor directly inhibit the B-ketothlolase or enoyl-CoA dehydrogenase sreps, nor does it affect the actir,rty of the carnitlne acyltransferases. The figure below shows the normal transport of electrons from FADH2 ro coenzlrne Q when the FADH2 is generated by the acyl-CoA dehydrogenases.

transfer flavoprotein (ETF), which then transfers rhe electrons to coenzyrne Q (via the ETF:Coe oxidoreductase,l

HH
-v-vPalmitoloyl CoA

FAD Acyl CoA DH

FAD (2H) Acyl CoA DH

flax, wheat, and canola). Monounsaturated oils need. a warmer ciimate, but not as warm as the tropics (o1ive, peanut). Knowing where a plant grows gives a iarge clue
whether the oi1 will be saturated, monounsaturated, or po1)-unsaturated. The diflerence in oil content between plants appears to be an evolutionary process. Kansas soil is very rich and supports growth of most plants.
as to

rAD (2H)
ETF

FAD ETF

FAD

FAD (2H)

ETF. OO

ETF. QO

The answer is C: acyl-(oA dehydrogenase. The acylCoA dehydrogenases catalyze the first step of the fatty acid oxidation spiral in that these enz),.rnes create a

CoQH2

CoQ

Electrontransport chain

Fatty Acid N4elabrri-.::-.

I 13
,.-,

ft

Tle answer is E: Medium chain acyl-CoA dehydrogenase. -:: child has MCAD (medium-chain acyl-CoA dehy-:--qenase) deficiency, an inability to completely oxi--:: fatty acids to carbon dioxide and water. With an

actlvated. This also contributes to the reduce

: :

neogenesis observed in patients with MCAD de -::--The answer is E: Carnitine

r.:--iD deficiency, gluconeogenesis

is impaired due to a

ol energy from fatty acid oxidation, and an inabll:o lul1y activate pyruvate carboxylase, as acetyl-CoA :-:i\-ates pyruvate carboxylase, and acetyl-CoA pro..-.'

,,:i

transporter. The chilc :-:.. ,. mutaiion in the enzyme which transports carnitrne -:--liver and muscle ce11s, leadlng to a primari- carr,r.-:. deficiency The carnitine stays ln the blood (rvhrch ie...:s
to its accumulation in the circulation). Slnce the lir e: carnitlne deficient, ketone body production ls minin:. at al1 times, even during a fast (thus, the lack ol berseline ketone bodies in the circulatlon under these condrtions). Fatty acids will rise in circulation, as they cannr.:
be stored in the cells as acyl-CoA. The muscle and iir-e: show er..idence of elevated triglycerides due to trigl,vceride s1,r-rthesis as the acyl-CoA cannot be degraded, anc acyl-CoA accumulates withln the cytoplasm. A defeci in carnitine acyl transferase (either 1 or 2) would not lead to elevated levels of carnitine in the crrculatron. nor would a defect ln MCAD (medlum-chain acyl-Co-t dehydrogenase). A defect in hormone sensitive lipase

'-.:tlon from fatty acid oxidation is greatly reduced.

:- an aitempt to generate more energy, medium-chai.n

.':ir- acids are oxidized at the trt ends to generate :: dicarboxylic acids seen in the question (see the -:ure below for an overview of ro oxidation). The -rding of such metabolites (dicarboxylic acids) in -:e blood is diagnostic for MCAD deficiency if there
ere mutations in any aspect of carnitine metabolism, ,eere would be no oxidation of fatty acids (the fatty ::rds would not be able to enter the mitochondria), :nd the dicarboxylic aclds (which are byproducts of -atty acid metabollsm) would not be observed. Simi-arly, a mutation in the fatty acyl-CoA synthetase (the actlvating etzyll;'e, converting a free fatty acid to an acyl-CoA) would also result in a lack of fatty acid oxidation, as fatty acids are not able to enter the mitochondria in their free (nonactivated) form.
-,,.

would show a decrease in free fatty acid

than the increase observed in the patient.

1eve1s,

rather

cH3-(cH2)n-c-o(r)
I

il

o il HO- CH2- (CHr,-C-O-

acyl-carnitine. Primary carnitlne deficiency is a lack of carnitine within the cell (such as a mutation in the carnitine transporter); secondary carnitine deficiency occurs when the carnitjne is sequestered in the form of acyl-carnitine (the carnitine cannot be removed from the acyi group, such as a defect in carnitine acyl transferase 2). Thus, elevated levels of acylcarnitine would be expected in a secondary carnltine defi.clency, but not in a primary carnitine deficlency In both types of carnitine deficiencies, fatty acid oxrdation is signifi.cantly reduced, so the levels of ketone bodies, glucose, lactate, and fatty acids would be similar under both condltions.
The answer is B: The answer is C: cisA9 Cl8:1. An lS-carbon fatty acid will generate an additionai acetyl-CoA, one NADH, and one FADH2 as compared to a l6-carbon fatty acid. Thus, the addition of two carbons will add 14 additional ATP

ilil -o-c-(cH2),-c-o-

The answer is C: lnsufficient energy for gluconeogenesis. Defects in fatty acid oxidation deprive the liver of energy

when fatty acids are the major energy source (such as during exercise, or a fast). Because ofthis, there is insuff,cient energy to s)'rrthesize glucose from gluconeogenic precursors (it requires 6 moles of AfP to convert 2 moles of p1'ruvate to I mole of glucose). Ac1'1-carnitines and dicarboxylic acids have no effect on the enz),rnes of gluconeogenesis, nor do they hinder the ability of the red blood cell to utilize glucose through the glycolytic pathway. Additionally, acetyl-CoA 1evels are low due to the lack of complete fatty acld oxidation and pyruvate carboxyiase, a key gluconeogenic enzqe, rs not fuliy

to the overall energy yield (10 AfP per acetyl-CoA,2.5 for NADH, and 1.5 for FADH2). An unsaturation at an odd carbon position will require the use of an isomerase during oxidation, and this will result in the loss of generation of 1 FADH2; an unsaturation at an even carbon position will require the use of the 2,4 dienoyl-CoA reductase, and this wiil result in the ioss of generation of 1 NADPH. Thus, an unsaturation at an odd position resuits in the loss of i.5 ATP while an unsaturalion at an even position results in the loss of 2.5 AfP Thus, in comparing two l8-carbon fatty acids, one with an unsaturatlon at position 9, and the other at position 6, the fatty acid with the double bond at position 9 will

LL4
A

Chapter 13 B

matrix
,,
-

Mitochondrial

P ) CHs\nn / CH2- CH2- CH2- C- SCoA Fatty ,' ltotat c=n1 acyl CoA | 7'--FAD i acvr coA i denydrogenas- f--t rno QH\ + -1.5 ATP ^ i
?

"&cl'"
B oxidation (thiee spirals)

]r"-or,ci*L12 tscoA

F+
+

s Acetyl CoA

B? CH3pn n rCH2-CH=CH-C- SCoA

@ wl/
enoyl

transL2Fally
enoyl CoA

4":o

43

,,,
ll,

\scoe

crEA3, crb-A6

l.-I
I

a,o

rsomerase

enoyl CoA ll

hvdratase 'l

CoA

trans-L2, ci*LG
O

Spiral

--J

CH3p1r17CHrl cH-

OH Br

CH2

- C- SCoA L-B-Hydroxy
acYICoA

l-'- Naoo-!1!1oy acylc'o lt-- NADH + H* + oenyorogenase @

Bil CH3p1r17CHr- C-

One spiral of B oxidation and the first step of the second spiral

Acetyl CoA

oo
CH,

-2.5 ATp

,l,^r1,A,".ij""o
2,4-dienoyl

54

tran*L2, cis-La

[ - C-

SCoA

B-Keto acyl CoA

;rz reduclase N,
CoA
J

NADPH + H+ ruROp*

l-'c"asa

-- CH3U,1 /CH2-C-SCoA + CHs-C- SCoA ltotal c=(n-2)l Fatty acyl CoA Acetyl CoA

?9

5 ,/\.,/\../\rZ\.,/

1^//O

4 2
con
I I

l'.

tscoA

trans-L3

enoyt

tsomerase

,4.,4'
B

53tz-O

.,/\,/c.' tscoA 4 2
I

tran*L2

oxioation

(four spirals)

Y 5 Acetyl CoA

Answer 9: Panel A: The steps of B-oxidation. The four steps are repeated until an even-chain fatty acidis completely converted to acetyl-CoA. The FAD(2H) and NADH are reoxidized by the electron-transport chain, producing ArP panel B: T-he additional reactions required for the oxidation of unsaturated fatty acids. The two new enz)'rnes required are the enoyl-CoA
isomerase and the

2,4 dtenoyl-CoA reductase.

1,re1d

one more ATp than the fatty acid with rhe unsaru_

ratlon at position 6. An overview of the fatty acid oxi_ dation spiral is shown above, along with the reactions
required for the oxidation ofunsaturated fatty acids.

oxidation will increase. If malonyl-CoA decarboxylase were inactivated, malonyl-CoA levels wouid remain elevated, and fatty acid oxidation would be inhibited.

oxidation is reduced, and as the levels decrease, fatty acid

ffi

An inacrivarlng mutation in acety'-CoA carboxylase would lead to an inabiiity to produce malonyl-CoA, which regulates fatty

ffr"

ansuuer is E: acetyl-CoA carboxylase

2.

acid oxidation through an inhibition of camitine acyl transferase 1. As malonyl-CoA levels increase, fatty acid

Inactivating mutations in either camitine acyltransferase I or 2 would lead to an inability to oxidize fatty acids, as they would not enter the mitochondria. A defect in medi_ um-chaln acyl-CoA dehydrogenase (MCAD) would also result in reduced fatty acid oxidation, as the initial step of the oxidation spiral would be inhibited once the fari

Famy Acid

Metabolism I

l5

)oA ,al2

acld had been reduced to about l0 carbons in length. The reactions catalyzed by malonyl-CoA decarboxylase
and acetyl-CoA carboxylase are shown be1ow.

o
CH.il
C

SCoA

Acetyl CoA
16

Biotin
Malonyl CoA decarboxylase

co. -l zATP ' \

Y

produce acetyl-CoA and oxaioacetate. The oxalLra..:-..:. is recycled to pyruvate, producing NADPH in the ::--cess, which is also required for fatty acld biosrnihe.-. A defect ln either carnitrne ac14 transferase wr1l not atie:: fatty acid biosy,r-rthesis, as those enz)rynes are require; to transport the fatty acid into the mitochondria lor ii. oxidatron. A lack of glucose-6-phosphate dehl,drogenase will not interfere wrth fatty acid syrthesis, as malic enzyme can pror-ide sufficient NADPH for the pathu ar MCAD is lnvoived in fatty acid oxidation and does nor
affect fatty acid sprthesis.

acetyl CoA arboxyrase

lt I

.OOO *

",

- C- CHr-

oo13 ilil
C

SCoA

Malonyl CoA

The answer is C:

19.

g-oxidation leads to the oxidation oi the cr-carbon of a branched chain fatty acid to generate an o,-keto acid, which undergoes oxidative decarboxylation. Thls reorients the methyl groups on the branched chain fatty acid such that they are on the
The answer is

B.

Acetoacetate will react with succi-

nyl-CoA to produce acetoacetyl-CoA and succinate (this costs I GTB as the succinate thiokinase step is skipped). The acetoacetyl-CoA is converted to two acetyl-CoA, each of which can generate 10 ATP when completely oxidized (each acetyl-CoA generates 1 GTf; 3 NADH, and 1 FADH2). The sum, then, is 20 minus the 1 iost in the CoA transferase step, for a net yield of 19 ATP

o,-carbon, rather than the B-carbon. ln this manner, the

translocase. Citrate translocase is required for citrate to exit the mitochondria and enter
The answer is C: Citrate

methyl groups do not interfere with the B-oxidation spiral (if the methyl group were on the B-carbon, a carbonyl group would be unable to form on that carbon, u,hich would block further oxidation of the fatt,r, acid). Answer choices A, C, D, and E are elimrnated as requirlng u-oxidation because, after one round of normal B-oxidation, the methyl group (or butyl group) will be on the g-carbon and would not interfere with the B-oxidation spirai. An overvlew of cr-oxidation is
shown be1ow.

the cytoplasm in order to deliver acetyl-CoA for fatty acld biosprthesis (see the figure below). Acetyl-CoA, which is produced exclusively in the mltochondria, has no direct path through the inner mltochondrial membrane. However, under conditions conducive to fatty
acid biospithesis (high energy levels, and allosteric inhrr rl

bition of the TCA cycle), citrate will accumulate and leave the mitochondria (see the figure below). Once

.\
\

a-Oxidation

in the cytoplasm, citrate lyase will cleave the crtrate to

CoA
Glucose

rida-

1", Ir
F
I

Pyruvate

The figure depicts the oxidation of phytanic acid. A peroxisomal cr-hydroxyiase oxidizes the s-carbon, and its subsequent oxidation to a carboxyl group releases the carboxyl group as carbon dioxide. Subsequent spirals of peroxisomal B-oxldation alternately release propionyl and acetyl-CoA.

acld
,1ase

naln
ited.

:I
S.

SC

OAA

i ,\ r'\
v

l"^,.ffiu:;
Acetyl CoA OAA Acetyl CoA

r{4.:

AS

iedtalso

..'. Citrate -.:---

-.-'-.

-..-'

ATP

step

Cltrate leaving the mitochondria and delivenng acetyl-CoA

c,vtoplasm for fatty acld sgrthesis.

lo

the

latty

The answer is A: Oxidation of very long chain fatty acids. The child has Zellweger's slmdrome, an absence of peroxlsomal enzyme activity Of the pathways listed as answers, only the oxidatlon of very long chain fatty aclds is a peroxisomal function. Fatty acid sprthesis occurs in the cytoplasm. Acetyl-CoA oxidation takes piace in the mitochondria. Glucose oxidation is a combination of glycolysis (cytoplasm) and the TCA cycle (mrtochondria). Triglyceride s1,r-ithesis occurs in the cytoplasm.

I6

Chapter 13
synthesis. Thromboxane 42 release from platelets is an essential eie_ ment of forming blood clots, and aspirin will block pros_

,.f5: The answer is C: To reduce thromboxane

It is the thromboxane inhibition whlch reduces the risk of blood clots. Leukotrienes and lipoxins require thr
en.zpe lipoxygenase, which is not inhibited by aspinr: These pathways arc outlined below.

taglandin, prostacyclin, and thromboxane s),nthesis.

Arachidonic acid
Cyclooxygenase

Cytochrome P45O

PGG2

HPETE

Epoxides

/
Prostaglandins

/\
Thromboxanes

Leukol.rienes

HETE

Lipoxins

diHETE

/\ /\

HETE

t5 .

nitine acyi-transferase 2 deficiency, the fatty acids are added to carnitine, but the acyl-carnitine cannot release the acyl group within the mitochondria. This will 1ead to an accumulatlon of acylcarnitine, which will lead to an
accumulation in the circulation. The end result of either deficiency is a lack of fatty acid oxldation, such that ketone body leveis would be minlmal under both condi_ tions, and blood glucose levels would also be similar in either condition. Insulin release is not affected by either deficiency, and carnitine levels, normally low, would not be significantly modified in elther deficienc;r

The answer is D: acyl-carnitine. With a carnitine defi_ ciency, fatty acids cannot be added to carnitine, and acyl-carnitine would not be sl.nthesized. With a car_

compared to belng covalently bound to proteins). The formation of malonyl-CoA, via acetyl-CoA carboxylase. requires biotin as a required cofacror (see the figure below). Citrate lyase, malic enzqe, acetyl transacy'ase (an activity of fatty acid slmthase) and acyl carrier protein (another component of fatty acid sl,,nthase) do not require bior in [or rheir acrivity.

o
cH3- 3 - scoA
Acetyl CoA

t7 The answer is C: C0I(-2 is specifically induced during

is induced durlng inflammatory conditions, whiie COX-1 is constirutively expressed.

ncetyt CoA carboxyrase

.o, _J,_oro Biotin f I II

inflammation. COX-2

l-*RoP * P,

Thus, when an injury occurs, and an immune response

is mounted at the site of injury, COX-2 is induced in those cells to produce second messengers that play a role in mediating the pain response. Specificaliy inhibiting the COX-2 isoz),rne will block the production of
those second messengers, without affecting the normal function of COX-I. lnhibiting COX-I may reduce the

ilil -O-C-CH2-C-SCoA
Malonyl CoA

oo

ls

frequency of heart auacks, and inhibiting COX-2 will block prostaglandin production via the cylco-oxygenase. Recent data suggests that certain drugs that specifically block COX-2 have unwanred side effects, such as an lncrease in heart attacks.

The answer is B: Activation of pyruvate carboxylase. Fatty acid oxidation increases the levels of acetyl-CoA

within the mitochondrial matrix, and acetyl-CoA is

neogenic
enzl.,Trre

potent activator of pyruvate carboxylase, a key gluco_

convert pyruvate to oxaloac_ to blpass the irreversible pyruvate kinase reaction). Acetyl-CoA cannot be used to s).nthesize net glucose, so it is not an effective precur_ sor of glucose production. Acetyl-CoA does not activate PEP carboxykinase (that enzyme is transcrlptionally controlled), nor does it affect pyruvate kinase (a cyto_ plasmic en4,rne). PFK-2 is nor regulated by acetyl_toA (phos^phorylation by protein kinase A is the key iegula_ tor effect for PFK-2 in the liver).
w111

(it

etate, a necessary first step

t&

The answer is E: Reduced ability to form malonylCoA. Biotinidase is required to remove covalentlybound biotin from proteins, which is how most of the biotin in our diet is received. In rhe absen ce of bio_ tinidase, individuals can become functionally biotin_ deficient, due to the iack of free biotin in the body (as

Fatty Acid
The answer is D: Fatty acid transport into the mitochondria. The man had eaten the unripe fruit of the ackee

Metabolism

ll7

the affected individual, leading to severe hypoglycemia.

Hlpoglycin has no effect on fatty acld

release from the

tree (from Jamaica). The unripened fruit contains the toxin hypoglycin A, which will interfere wlth carnitine's ability to transport acyl-carnitine groups across the lnner mitochondrial membrane. This leads to a complete shutdown of fatty acld oxidation in all tissues in

adipocyte, or fatty acid entry into liver cells. Fatty acid oxidation is not directly lnhiblted, nor does this toxin directly inhibit the complexes of the electron transport chain and the proton-translocating AfPase.

Chapter 14

X-{&P Shux?E

ercd Gxidative Reacti*xas

This chapter toyers qutstians relsted to the pclttos( phosphate shunt pathway and rea$ions that generarc dnd prote# againsi ra.dical axygttl sptcies. Intey*ctiotts oJ' rhls pathtlay with oth* metabolic Tsatl*,v ay s sre crlsa emphssized.
He dres thar night in his s1eep. Thls is due ro u,hich of the foilowlng? (A) Ethanol stimulating barbiturare absorplion b1, the
stomach

QUESTIONS
Select the single best answer.

(B) Ethanol inhjbirion ol a cl,tochrome p450 s),slem (C) Acetaldehyde reacting i,vrth the drr_rg, crearing
toxic compound

(D) Acetyl-CoA producrion

leads to enhanced encrgv

pro,

A 52-year-old man has had bor-rts of alcohol abuse tn his past. During his binges, he rakes aceiamrnophen to help control some muscle pain. He then gers very
r11

(E)

ducrlon, u.hrch slnergizes lvirh barbirurate acrion Ethanols dehydration eifecr leads lo roxic concen_ tratlons ol rhe seizure medrcation in the blood

(nausea, r,omiting, and right upper quadrant pain),

and ls rushed to the emergency department. A potenrial treatmenr ior this patients svmptoms is to rake whrch of

the foilowing? (A I Asplnn (B) A mercapran

A chronic alcoholic presents to the emergency department r,vith n,vstagmus, peripheral edema, pulmonary edema, ataxia, and mental confusion. The physician orders a test

to determine if there is a ritamtn deficrencri An enzvme used lor such a resr can be w.hich oi the foliou.ing?

(C) Rrfampin (D) Iron (E) Vitamin C

A34-year,o1d man was prescribecl barbrturates 6 months ago lor a seizure disorder. Hor,vever, wlth time, the phy_

(A) Transaldolase (B) Aldolase (C) Tiansketolase (D) B-ketothiolase (E) Acet),lcholine sp-rthase
researcher ls studl,ing the HMp shunt pathrval- in extracts ol red blood ceils, in the absence ol NADpr, and in whlch PFK-1 has been chemica111, inactivated. Which

sician has had to increase his daily dosage to maintain the same therapeutic drug level. This is due to whlch ol the follolving?

(A) Downregulation of drug receptors on the ceil surface (B) Decreased absorprion ol the drug lrom rhe sromach (C) Increased sl,nthesis of opposLng neurorransmitrers (D) Indr-rction of drug-merabolizing enzymes (E) Induction ol targeted enzyme synthesis

carbon substrates are required

phosphate in rhis system?

to

generate ribose_5,

(A) Glucose-6-phosphate and sedoheprulose-7-phosphate

(B) Glucose-6-phosphate and glyceraldehyde-3-phosphate (C) Fructose-6-phosphare and g1yceralc1eh1,de-3-phosphate

Considering the patient in question 2, one night, the patient consumes a large amount ol alcohol. He
continues to take his usual dose of seizure medication.

(D) Fructose-6-phosphate and pyruvate

(E) Glucose-6-phosphate and p),ruvare

rt8

HMP Shunt and Oxidatrve Reactions

I 19

Whlch one of the following is an obligatory intermediate ln the conversion of ribose-5-phosphate to glucose6-phosphate?

(A) Pyruvate (B) 1,3-bisphosphoglycerate (C) Oxaloacetate (D) Xylulose-5-phosphate (E) 6-phosphogluconate

(A) (B) (C) (D) (E)

1t

Superoxide

Nitrogen dioxide Nitrous oxide Hydrogen peroxide Peroqmitrate

ma1e,

A 25-year-old African American

in good health.

23-year-oid man

of

Medrterranean descent was

recently prescribed ciprofloxacin to treat a urlnary tract infection. After 2 days on the drug, the patient was feeiing worse, and weak, and went to the emergency department. He r'vas found to have hemolytic anemra. This mosr likely resulted due to w-hich of the following? (A) lnduction of red blood cell cytochrome P450s, leading to membrane damage (B) Oxidative damage to red blood ce11 membranes

had read about fava beans in "Silence of the Lambs. For dinner one night, the man had liver with fava beans and a nice Chianti. About 8h after eating the beans, the man was tired and weak. Blood work showed hemolytic anemia. This patient most likely has a defect in regeneraring which of the following?

(A) NADH (B) NAD. (C) Reduced glutathione (D) Oxidized glutathione (E) ATP

(C) Drug induced ion pores

membrane

in the red blood ceil

l2 A 52-year-old male
complarned of sudden onset of left-

(D) Drug induced inhibition of the HMP shunt pathway (E) Oxidative damage to bone marrow, interferlng with
red blood celI product ion

sided chest pain radiating down his left arm. Rapid

breathlng, sweating, and a feeling of doom accompanied this. He was rushed to the emergency department. An angiogram revealed 90% occlusion ol the left anterior descending artery (lAD) and no occlusions in any other artery The IAD was opened by angioplasty However, shortly after this procedure, with normal blood flow through the lAD, the patients conditlon worsened. This was most likely due to which of the following? (A) Disruption of the HMP shunt rn cardiac cells (B) Damage to healthy ce1ls by loss of essentlal enzymes from cel1s due to membrane damage (C) Development of intimal narrowing in another artery

You are seeing a male patient of Afncan American

descent, whose grandparents live in a chloroqui.ne resistant malaria belt in Africa. He wants to \'1s11 his grandparenls, and you want to give him primaquine as a malaria prophylaxis, but before you do so, you should test the patient for which of the following nonsymptomatic enzlmaiic defi ciencles?

(A) Tiansketolase (B) Pyruvate dehydrogenase (C) o-Ketoglutarate dehydrogenase (D) Glucose-6-phosphate dehydrogenase (E) Glyceraldehyde-3-phosphate dehydrogenase
A patient has an insldlous and steadily progressing neurologic disorder thal, after several years, results in wasting and paralysis of the muscles of the limbs and trunk, loss of abillty to speak, and swallowlng difficulties. His paternal uncle had the same disease. A mutation in which en4.me may lead to these sy'mptoms?
13

(D) Radical-rnduced damage once blood flow

reinitrated

was

(E) Lack of glycogen for AfP sprthesis in the heart

Consider an intestinal epithelial
ce11

in

S phase,

and for

which the major, active biosynthetic pathway is nucleotide sgrthesis. Which one of the following best represents the actlvity state of a series of key en4.mes under these conditions?

(A) Superoxide dismutase (B) Catalase (C) Myeloperoxidase (D) NO s).nthase (E) Tyrosine hydroxylase
researcher has generated a celi line in which the y-glutamyl cycle is defective, and glutathlone cannot be sl,rithesized. Which radical specles mrght you inltially expect to accumulate in thrs cell?

Glucose-

6-phosphate

dehydrogenase
(A)
(B)

Transketolase
Active lnactive Active

PFK.1

Active
Active
lnactrve lnactrve lnactrve

Active
lnactrve lnactrve

(c)
(D)
(E)

Active

Active

'a;!in'i:::::

120 14

Chapter lzl

A researcher is studying cultured human hepatocytes and ls examining the specific condition in which fatty
acid synthesis is activated, but the ceil remains in the

G" phase of the cell cycle. Under such conditions. what would be the activity state of the following
enzymes?

Glucose-6-phosphate dehydrogenase
(A)
(B)

Fructose-1,

Transketolase
Active Actrve
lnactrve
ACTiVe

Transaldolase
lnactive Active Active lnactive Active

6-bisphosphatase
lnactive Active lnactive Active lnactive

Active Active Active

lnactrve

(c)
(D)
(E)

lnactive

lnactive

t5 lndividuals with a superactive glutathione reductase will develop gout. Thls occurs due to which of the
following? (A) Activation of glucose-6-phosphate dehydrogenase (B) Inhibition of glucose-6-phosphate dehydrogenase (C) Activation of transketolase (D) Activation of transaldolase (E) Inhibition of transketolase
16

a healthier life style and replaced eggs and coffee for

breakfast with fruit.luices and whole-wheat toast. Within 2 weeks of changing his diet, the man developed severe muscle pain in his arms and shoulders. The muscle pain could be the result of whlch of the foilorvlng? (A) Induction of a detoxlfl,rng cytochrome P450 system

(B) lnhibrtion of a detoxifying cytochrome P'150 system (C) Increased mevalonate inhibiting actin/myosin interactlons

Glucose-6-phosphate labeled in carbon 6 wrth added to a test tube wrth the enzlrnes phosphohexose isomerase, PFK-1, a1do1ase, transketolase, and transaldolase. AfP was aiso added to the test tube. At equrlib-

1aC was

(D) lncreased mevalonate stabilizing actin/myosln interactions

(E) HMG-CoA stimulatron of calcium efflux from the

sarcoplasmic reticulum

num, in which position would the radioactive label be found in the newly produced ribose-5-phosphate?

t9 A patient is recoverlng from

(A) I (B) 2 (c) 3

(D) 4

acute respiratory distress sl.rLdrome (ARDS). Which of the follomng is a ma.1or

antioxidant found in the fluld lining the bronchial epitheiium needed ln high concentration for recovery
from ARDS? (A) Glucuronic acid

(E) t7

Which one of the following disorders would lead to

lncreased activity of the HMP shunt pathway? (A) Glycogen phosphorylase deficlency

(B) Pyruvate (C) Sorbitol (D) Glycogen (E) Glutathione

20

(B) (C) (D) (E)

Glucose-6-phosphatase deficiency Fructose-1,6-blsphosphatase deficiency Pyruvate kinase deficiency Pyruvate dehydrogenase deflciency hypercholes-

Which of the following biochemical pathways produces

the antioxidant referred to in the previous question? (A) TCA cycle

t8 A 45-year-old man rvas diagnosed with

terolemia, for which he was prescribed a statin. After a monih on medications, the patlent decided to adopt

(B) Glycolysis (C) y-Glutamyi cycle (D) HMP shunt (E) Polyol pathway
I

::;i:E
HMP Shunt and Oxidative Reactrons

l2l

{},iSWERS
I
lhe answer is B: A mercaptan. The man is suffering from acetaminophen poisoning. As shown below,
\,{EOS (the microsomal ethanol oxidlzing system, also

glutathlone to the compound for safe excrerion -,: * glutathione is a mercaptan (a compound riith :r i:::

sulfhydryl group). Individuais wrth acetaminsrpr.::.

poisoning are given N-acetyl cysteine as a mechan.s::l

named CYP2E1) will convert acetaminophen into a toxic intermediate. In a chronic alcoholic, the MEOS has been induced and ls very active. The loxic inrermediate (NAPQI) can be rendered inactive by adding
il H^ -N-C-CH3

to increase glutathione production. Iron and \rtain,:, C will not aid in detoxifylng the toxic inrermedia:. Rifampin blocks RNA polymerase in bactena. Asptrin will block cyclooxygenase, but wili not stlmulaie
NAPQI excretlon.

H-ilH-il -N-C-CH"

oo

Kidnev. a,A, ,rin.<-[l_rl to,',-no.*

Glucuronate

0 -***'* Q-sru
oH
I

'N-C-CH"

soa

Acetaminophen

EtoH--6-+l

CYP2E1

'.r-8 -cr, "i

<SG
OH Mercaptouric acid

N-acetyl cysteine

io

? N-C-CH3

II H^ 'N-C-CH3

GSH
Glutathione

s-transferase

Vo -Cilp.i".--

S-protein

I
Kidney, urine

NAPQI (N-acetyl-pbenzoquinoneim ine) (toxic intermediate)

The answer is D: lnduction of drug-metabolizing enzymes. Barbj.turates are xenobi.otics, and the body induces specific cytochrome P,i50 systems to help
detoxify and excrete the barbiturates. When the man
first begins taklng the drug,
a 1ow

concentration of drug

is sufficient to exert a physiologlcal effect, as the drug detoxifpng system has not yet been induced. As the
detoxifying system is induced, however, higher concen-

trations of drug are required to have the same effect, as the rate o[ excretion of the drug is increased as the detoxification system is induced. The "tolerance" to
drugs, Ln this case, is not due to downregulation of drug receptors or decreased absorption of the drug from the stomach. There is no lnduction of target gene expression, Ieading to enhanced drug action, nor are opposing neurotransmitters expressed. The tolerance comes about due to enhanced inacti.vation of the drug due to the induction of drug-metabolizing er.:zqes.
The answer is B: Ethanol inhibition of a cytochrome P450
';,.*:!,,

high levels of barbiturates being taken (due to the tolerance) are now toxic (the system that breaks down the drug has been lnhibited). Ethanol does not increase absorption of the drug from the digestive tract, nor does acetaldehyde, ethanol's oxidation product, react with barbiturates. Barbiturate action is not affected by energy production (acetyl-CoA). The ethanol inhibition of cytochromeP450 systems is also not due to ethanol's dehydration effect.

The answer is (: Transketolase. The patient is experiencing the symptoms of vitamin B, (thiamine) deficienqr

Ethanol blocks thiamlne absorption from the gut, so in the Unlted States, one will usuaily only see a B, deficiency in chronic alcoholics. One assay for B, deficiency is to measure transketolase activity (which requires B, as an essential cofactor) in the presence and absence of added B,. If the activity level increases when B, ls added, a vitamin deficiency is assumed. None of the other

system. Ethanol inhlbits the drug detoxifpng system for barbiturates; thus, in the presence of ethanol, the

enzqes listed (transaldolase, aldolase, B-ketothiolase, and acetylcholine syrithase) require B, as a cofactor,

122

Chapter t4 and, thus, could not be used as a measure of B,

1evels.

(note the breakage of a carbon-carbon bond, and then the syrrthesis of a carbon-carbon bond to generate the product of the reaction).

A reaction catalyzed by transketolase is shown beiow

occur. In additlon, PFK-1 has been made nonfunctional,

cH2oH
I

C=O

HO_C-H
I

H_C_OH
I

cH2oPO32-

Xylulose S-phosphate

HO \0
C

5-phosphate (X5tl step 1 in the figure below). The X5p will be epimerized to ribulose-5-phosphate (Ru5f; step 2 in the figure below), and then isomerlzed ro R5p (step 3 in the figure below). Glucose-6-phosphare can_ not be used as a substrate because it cannot be con_ verted to G3P (due to the block in pFK_l). pyruvdle cannot be used as a substrate in extracts of red blood cells because such cells do not have pyruvate carboxi. 1ase, so the pyruvate cannot be converted to either F6p or G3P

substrates will react, using transketolase as a substrate, to generare erythrose-4-phosphare (E4p) and xylulose_

such that glyceraldehyde-3-phosphate (G3p) cannot be produced from either fructose-6-phosphate (F6p) or glucose-6-phosphate (G6p) In order ro generare ribose-5-phosphare (R5p) under these coriditions, both F6P and G3p need ro be provided. These rwo

H-C-OH
H-C_OH H_C-OH
I I

J HtOUtOSe-5-P

cH2oP032Ribose s-phosphate
lntamtne

.I

l
I

PYroPhosPhate I

Transketolase

Nonoxidative reactions

HO \//
L,
I

Fructose-6-P

clyceraldehyde-3_p

H-C-OH
I

cH2oPO32G

lyceraldehyde 3-phosphate

.6

+
cH2OH
I

The ansurer is D: Xylulose-5-phosphate. In order for rjhose-5-phosphate to be converted to glucose_6_phos_

phate, the nonoxidative reactions of the HMp shunt ible reactions). In order for this to o..rr., the ribose_5_ 2 in the figure on page 123). R5p and X5p then initi_ figure on page 123) to generare fiuctose_6_pirosphate, which can be isomerized to giucose_6-phosphate (rt.p : in the figure on page 123). Glyceraldehyde_3_phosphate
pathway must be used (the oxidative steps are not revers-

U-LJ

HO-C-H I H-C-OH I H-C-OH H-C-OH

I I

phosphate is tsomerized to ribuiose_5_phosphate, which is then epimerized to xylulose-5-phosphate (steps 1 and

ate a series of reaoions utilizing transketolase (step 3 in the figure on page 723) and transaldoiase (step 4 in the

cH2oPOs2-

Sedoheptulose 7-phosphate

is also formed during this senes of reactions, which then goes back to fructose-6-phosphate production.
:',.}

The answer is C: Fructose-6-phosphate and glyceraldehyde-3-phosphate. In the absence of NADp*, the oxidative steps of the HMp shunt pathw ay are nonfunctional, so only the nonoxidatlve steps will

Pyruvate, oxaloacetate, 1,3-bisphosphoglycerate, and 6-phosphogluconoate are not obligatory intermediates

in this conversion.

HMP Shunt and Oxidative Reactior-3 Ribu lose-

lll

(Clisomerase
Xylulose-S-P

eprmerase

--

1.",

-z
A-

Ribose-S-P

Xylulose-5-P Nonoxidative reactions

,runan",o,ffi
Gtyceraldehyde -3-P

/ \

s"oor,"ptrto."-

,run.u,O",*"XD

__*E'.y*,"= Fructose-6-P Glyceraldehyde-3-P

Fructose-6-P

-/

\nosPnonero."i.oruru.9,/ \

9\ctucos*o*/

Dihydroxyacetone phosphate

It

The answer

damage to red blood cell glucose-6-phosphate dehydrogenase deficiency and is incapable of regen-

is B: Oxidative

membranes.

The man has

erating reduced glutathione to protect red blood

ce11

cooI

CHz
I

Glycine

HN

U-IJ
I

membranes from oxidative damage. In the presence o[ a strong oxidizrng agent (the new drug the paiienr was takrng), the red cell membranes undergo oxidatir-e damage and the red ce11 bursts, leading to hemolytic anemia. This is all due to a lack of protective glutathione in the membrane. As the red cell lacks a nucleus. the celi cannot induce new gene s).nthesis. The drug the patient was taking does not induce ion pores in red ceil membranes or inhibit the HMP shunt pathway It also does not cause oxidative damage to bone marrow. The

HS-CH2-CH
I

Cysteine

drugs to avoid whl1e prescrlbing for a patient with a G6PDH deficiency include prlmaquine, dapsone, nitrofurantoin, and sulfonylurea. The reduced (Panel A) and oxidrzed (Pane1 B) forms of glutathione are indicated to
the side.

HN

C=O
I

1l:1-tn:
a:>-

CHz
I

CHz
I

Glutamate

HCNHJ

a-r-

lch
;.ilii r tl-

cooB

I"

The answer is D: Glucose-5-phosphate dehydrogenase. Given the demographrcs of the patients ancestry (and

cooI

cooI

lrn
-L.
raie.

CHz
I

CHz
I

Glycine

HN
I

HN

:ni
1:.le

lch
i rr

II HC-CH,-S-S- CH,-CH t-l HN HN

C=O
I I I

L,-U

C=O

I1.

C=O
I

and

CHz
I

vt t2
I

CHz
I

CHz
I

Glutamate

HCNHd

HCNH]

the need for obtarning an accurate history), and the fact that the patient is a ma1e, the patient may have glucose-6-phosphate dehydrogenase deficiencl (an X-linked disorder). If a person with this enzyme deficiency is given primaquine, which is a strong oxldizing agent, hemolytic anemia is likely to develop. If a physiclan suspects that a patient may have such an enzymatic deficiency, it is imperative to check before prescriblng strong oxidizing agents to the patient, or prescribe another antlmalarial prophylaxis that is not a strong oxidizing agent (such as tetracycline). If individuals were deficient in transketolase, pyruvate dehydrogenase, u-ketoglutarate dehydrogenase, or
glyceraldehyde-3-phosphate dehydrogenase, red cell lysis would not occur. One should also recal1 that the red ce11s lack mitochondria, so these cells do not contain pyruvate dehydrogenase or s-ketoglutarate
dehydrogenase.

cooI

t" coo-

Answer 7: Panel A indicates reduced glutathione (GSH) while Panel

B indicates oxidized glutathrone (GSSG)

124 g

Chapter 14

is A: Superoxide dismutase. The patient is expenencing the sl.mptoms of familial ALS. A muta_ tion in superoxide dismurase I (SODI) in humans has
The answer
been linked ro the development of famiiial ALS through an unknown mechanism. Familial ALS only constitutes between 5ob and 10o/o of all ALS cases diagnosed. The disease process, u.hen SODI is mutated, is not linked to a loss of enzl,rnatic activlty, although the SODl may have been mutated such rhat it wrll produce other radical spe_ cies and is no longer specific for superoxide. A second model proposes a misfolding problem similar to pnon disease. For more information on such models see

in individuals wirh glucose-6-phosphare dehydrogenase deficiency; in indlviduals wirh a normal G6pDH, rhe
oxidizing agenr is handled by glutathione. The red blood cells, under these conditlons, do not have a problem in regeneraring NADH, NAD*, or Afp

12

reinitiated. The patrent is experienclng rschemlc reperfusion injuryr When oxygen delivery to cardiac ce11s was compromised, the electron transfer chain in the mitochondria was fully reduced, as the terminai oxygen acceptor was mlssing. When oxygen is reintro_
was duced to the
ce11,

The answer is D: Radical induced damage once blood flow

Nature Med. 2000;6:1320-132I and Ann Neurol. 2007 Dec;62(6):553-559. None of the other erq.mes listed
(catalase, myeloperoxidase, NO sy,nthase, and tl,rosine hydroxylase) have been linked to the development of ALS, or an ALSlike disease. The reactron catalyzed bv SODI is shown below.

at a hlgh concenrration, the likehhood

of electron transler from reduced coenzyme

gen is increased, such that the possibility of superoxide generation is increased. The superoxide produced reacts with lipids and proteins and can iead to cell death above

to oxy_

2
2H+

az-

Superoxide
Superoxide dismutase

thar origlnally occurring from the initial heart attack. Radicals do not form during the ischemrc event since oxygen ls missing from the tissues. There is no effect on glycogen srores Qr rhe HMp shunr pathway under these conditions. Intimal narrowing occurs over a long time period, not over'the short time period covered rn this case. lnjured ceils do leak enzymes into the blood_ stream, but these enzymes do not cause the death of
orher. healthy celis.

o2
HzOz Hydrogen peroxide

13

is E. Under the conditions described, DNA sy,rithesis is occurring without any requiremenr
The answer for NADPH (such as fatty acid synthesis). Under these

The answer is D: Hydrogen

of glutathione, rhe enzyme glutathione peroxidase wlll be less active due to the lowered concentratlon

peroxide. ln the absence

conditions, NADPH

1eve1s

are high and

glucose_6_

of glurathione. Glutathione peroxidase catalyzes the

oxidation of two reduced glutathione molecules by hydrogen peroxide, generarlng oxidized glutathione
and two molecules of water. As glutathione peroxidase

is one mechanism whereby hydrogen peroxide levels

are reduced, hydrogen peroxide would be expected to accumulate, and can then lead to radical damage of membrane proreins and liplds. Glutathione peroxidase does not require, or react with, superoxrde, nitrogen dioxlde, nirrous oxide, and peroxynitrare. It is possible that under these conditions, superoxide would also accumulate, due to the increase in concentration of one of the reaction products oI superoxide dismutase, hydrogen peroxide. However, there is no evldence that hydrogen peroxlde accumulation w111 inhlbir rhe reaction catalyzed by superoxide dismutase.

phosphate dehydrogenase is inactive. The cell requires ribose-5-phosphate, however, for nucleotide biosl.,nthe_ sis, and this is sprthesrzed from fructose_6_phosphate and glyceraldehyde-3-phosphate using the nonoxida_ tive reactions of the pathway Thus, both transketolase and transaldolase will be actrve under these conditions. PFK-1 is actlye as we11, as the only way to generate glyc_ eraldehyde-3-phosphate from a sugar precursor is via enzymes of the glycolytic pathway

l/t

The answer is B. The conditions of the cell indicate that NADPH is required for fatty acid synthesis, but there is no need for ribose-5-phosphate, as the cells are in Go phase and are not undergoing DNA synthesis
(so nucleotides are not requlred). The HMp shunt will utilize the oxidative reaclions to generate NADPH, and then the ribulose-5-phosphate produced will use the nonoxidatlve reactions to regenerale glucose_6 -phosphate. For this to occur, transketolase, transal_

fi

The answer is C: Reduced glutathione. The patient has glucose-6-phosphare dehydrogenase deficiency, and his red blood cells cannor convert oxidized glutathione to reduced glutathione due ro a lack of NADPH.

dolase, glucose-6-phosphare dehydrogenase (as the

Fava beans contain a potent oxidizing agent that wil1, in some paiienrs (but not all), lead to hemoiytic anemia

major oxidative enzyme of the pathway), and fruc_ tose-1,6-bisphosphatase all have to be active. These
nonoxidative reacrions are indicated in the figure on
page 125.

f-

HMP Shunt and Oxidative Reacrions

rogenase

125

'DH, the
blood rblem in
ed

3 Ribulose-S-P

Xylulose-S-P ood flow

r_

z,z
A-

Ribose-S-p

Xylulose-5-P

,rrn.na,o,)fu
Gtyceratdehyde -s-p

schemic cardiac

:hain in
.erminal

/ \ \

r"oon"o,rtose-7-p
Erythrose-4-p

transatoorase)b

reintroelihood to oxyeroxide
d reacts
h above

-/

attack. rt since

Fructose 1,6-bisphosphate

Answer 14

c effect ' under

a long

15. The answer is A: Activation of

glucose-6-phosphate

utiiize NADPH and reduce oxldized glutathione to reduced glu,

tathione, generaring NADP*. If Glutathione reducrase is superactive, NADP+ Ievels accumuiate, which activates glucose-6-phosphate dehydrogenase. This will lead to

dehydrogenase. Glutarhione reducrase

will

ered in

blood3ath of

NADPH production via the oxldative reactions of the HMP shunt, aiong with ribulose-5-phosphate (Ru5p).
:nbed,
-ement

3rd position). Tiansketolase wiil allow these two compounds to exchange carbons, which would generare
erythrose-4-phosphate (E4B labeled in the 4th posirion) and xylulose-5-phosphate (X5f; labeled in the 5th posi_

position \.irh r4C) musr pass through glycolysis ro pro_ duce fructose-6-phosphate (F6p, labeled in the 6th posi_ tion) and glyceraldehyde-3-phosphate (labeled in the

In order for the nonoxidative reacilons [o occur, the glucose-6-phosphate (G6B iabeled in the 6th
take p1ace.

: ihese
ose-6quires

The Ru5P w11i lead to increased ribose-5-phosphate pro_ duction, increased 5'-phosphonbosyl 1'-pyrophosphate (PRPP) production, and increased 5,-phosphoribosyl

tion). The X5P can then go to ribulose-5-phosphate

(Ru5P) and ribose-5-phosphate (R5p), labeled in the fifth positions. The E4p (labeled in the 4th position) can react with anorher molecule of F6p (1abe1ed in rhe 6th position) uslng transaldolase to generate sedoheptulose 7-phosphate (Se7B labeled in rhe 7th position) and glyceraidehyde-3-phosphate (G3p), labeled in the 3rd position. Tiansketolase will then convert the SeTp and G3P to R5P and X5f; both labeled in the 5th posrrions. The nonoxidatlve reactions can be seen, schematically, in the figure below

!nthe;phate cxlda:iolase

1'-amine levels. This eventually leads to increased purine production, in excess of what is required. The excess purines are converted to uric acid, and excess uric acid will lead ro gout. A superactlve glutathione reductase wiil not lead to an alteration tn the actrvities
of transketolase or transaldolase

Itions.
:

glycis via

t6

5. Given the enzymes present, only the nonoxidative reactions of the HMp shunt would
The answer is E:

licate

I
"r*",u""

3 Ribulose-S-P

;, but
cells
.hesis
r

a|.",",;
Hibose.S.P

eprmerase

will

'' ;"::",*x;
Grycerardehyde

Xylulose-5-P

DPH,

ll use
:se-6 nsal-

a, 4 \

Sedoheptutose-7-p

transatoot"se)b
Erythrose-4-P

; rhe
fruc'hese 'e on

"ffi:;;ffiz
Fructose-6-P

-/

Fructose 1,6-bisphosphate

6,,:;,Ji-.:::",,.,"

:,:,

:.: I

126 17

Chapter 14

The answer

is B:

Gtucose.6-phosphatase deficiency.

The HMP shunt can have increased actrvity under two conditions, one being an rncrease in the cofactor NADp. levels and the other being an increase in the substrate levels (glucose-6-phosphate) The only enzyme listed, which when defective would lead to an increase in either glucose-6-phosphate or NADpH, is glucose_6_phos_
phatase. A deficiency rn g11.c6gsn phosphorylase would not produce glucose-1-phosphate; thus, there wouid not be an increase in the HMp shunt under these conditrons. A deficiency in frucrose-1,6-bisphosphatase deficiency would impair gluconeogenesis and would not lead to the synthesis of glucose-6-phosphate. Deficiencles in either pyruvate carboxylase or pyruvate dehydrogenase would lead to pyruyate accumulation and NAD. accumulation, but not NADp. or glucose-6-phosphate accumulation.

increasing cannot be correct; they are reduced presence of a statin. Statins do noi bnng about calcium elflux lrom the sarcoplasmic reticulum.
ler.e1s are

in the

19

18

P450

side effects of statins, namely, muscle damage and pain. This may occur due to an inhibition of coen4.me Q sp-rthesrs (which requires a product clerited lrom mevalonic acid) and a lack ol energy generation in the muscle. The reason this comes about rs rhat statins are detoxified through a cyrochrome p45O sysrem, and the particular sysrem that works on srarins is inhibited by
absence of the juice, due to the decreased rate of destruction. The artificiallyinduced higher levels of statins then lead to muscle damage. Statins inhibit the conversion of HMG-CoA ro mevalonic acid. (catalyzed by HMG_CoA reductase). Thus, answers indicating that mevalonrc acid

The answer is B: lnhibition of a detoxifying cytochrome system. The patient is suffenng fron rhe porenrial

The answer is E: Glutathione. Glutathione is the ma.lor antioxidant in the fluid lining the bronchlal eprthelium. It is essential for recovery of these tlssues. Depletion ol glutathione in the airway is thought ro g."atli increase a person's susceptibiljt)- ro upper respirarory infec_ tions such as influenza. Glutathione is formed in the y-glutamyl parhway, and oxidrzed glurarhione is regen_ erated ro reduced glutathione using NADpH produced by the HMP shunt parhwa;,r None of the othei answers (glycogen, sorbitol, pyruvare, and glucuronate) offer protection against oxidative damage. G11.cogen is utilized for the storage ofglucose. pyruyare is the eid producr ol glycolysis. Sorbitol is a producr of the polyoi parhwa)l Glucuronic acid is used for xenobiotic metabolism. in general, to increase the solubility of the xenobiotic and to prepare it for excretion. The answer is C: y-Glutamyl
duced_

20

cycle. Glutathtone is pro_ via the y-gluramy1 cycle; the HMp shunt pathwal

in the

grapefruir.luice. Thus, in the presence of grapefrult.lri.., the effective intracellular levels ol statins are highei rhan

provides the NADpH that allows oxidized gluiathione to be converted to reduced glurathione. The other path_ ways listed (TCA cycle, glycolysis, HMp shunt, and the polyol pathway) do not pronde for glurathione rynrhe_ sis. The TCA cycle is designed to oxidize acetyl_CoA to carbon dloxide and water. Glycolysis rs the entiy point oi sugars lnto metabohsm. The HMp shunt pathway gen_ erates five-carbon sugars and NADpH, and the po1yo1
pathway generares sugar a1cohols. The y_glutamyi cycle is shown in the figure below

ADP +

P;

ATP

ii t
1
I

Amino acid

O
y-Glutamyl
transpeptidase

Glycine

ADP +
Cysteine ATP

P,

Answer 20: In cells of the intestine and kidney amino

acids can be transported across the cell membrane by reacting wlth glutathione to form a y_gluLamyl amino acid. I he amino acid is released inro the cell and glu_ tathione is resltrthesized. Hower.er, the ma.lor role ol this cycle is glutathione slnthesis because many tissues

Amino
acid

ATP

ADP + p;

y-glutamylcysterne and then the condensatron of y-glutamylcysteine with glycine to lorm glutathrone.

lack the transpepriclase and 5-oxoprolinase activities. Thus, the reacrrons performed by most cells include the condensation of cysteine and glutamate to lorm

' I

I I

: :

I I :

:: t:

::l

Chapter 1 5

Amino Acid Metabolism

and the Urea Cycle
lhis chapter quiTTes the student on amino
-v
acid
most often results from a defect in which of the following
en4..rnes?

tob olism and pr o ducts denv e d fr om amino acids.

QUESTIONS
-lect
the single best answer.

Routine newborn screening identified a child with elevated 1evels of phenylpyruvate and phenyllactate in the blood. Despite treating the child with a restricted
diet, evidence of developmental delay became apparent. Supplementation with which of the following would be beneficial to the child?

(A) (B) (C) (D) (E)

Phenylalanine hydroxylase

NADPH oxidase Dihydrofolate reductase

Tyrosinase

Homogentisic acj.d oxidase

after

A newborn becomes lethargic and drowsy 24h

(A) Tyrosine

birth. Blood analysis shows hyperammonemia, coupled with orotic aciduria. This individual has an enzyme deficiency that leads to an inability to directly produce which of the following?

(B) 5-hydroxytr'lptophan (C) Melanin (D) Phenylalanlne (E) Alanine

(A) Carbamoyl phosphate (B) Ornithine (C) Cltrulline (D) Argininosuccinate (E) Arginine
Considering the patient in question 3, orotic acid levels in this patient due to which of the following? (A) Elevated ammonia (B) Elevated glutamine

A newborn has milky white skln, white hair, and redappearing eye coior (see the figure below). This disorder
are high

(C) Bypassing carbamoyl phosphate s;mthetase II

(cPS-II)

(D) Bypassing aspartate transcarbamoylase (E) Inhibrtion of carbamoyl phosphate slmthetase I

(cPS-I) Considerlng the patient discussed in the last two questi.ons, a potential treatment for the patient is supplementation with which of the following? 1A) Arginine and glutamine (B) Lysine and glutamine (C) Arginine and benzoate

(D) Lysine and benzoate

(E) Glutamine and phenylbutyrate

t27

r,"=

128
'.f,

Chapter 15
Parents bring their 6-year-old son to the pediatrician due to the parents being concerned about "mental retardation." Blood work demonstrated a microcytic anemia

(reminiscent of Marfan sl.ndrome patlents), scoliosr:

pectus excavatum, displaced lens, and muscular hlpctonia. Blood work is likely to show an elevation of whicl of the following metabolites?

and basophilic stippling. During the patlent history became apparent that the boy often stayed with his grandparents, who owned a I5O-year-old apartment. The boy admitted to eating paint chips from the radiators in the apartment. The boy's anemla is most likely the result of which one of the following? (A) lnhibition of lron transport

it

(A) Methionine (B) Phenylpyuvate (C) Cysteine (D) Fibrillin fragments (E) Homocystine
11

(B) (C) (D) (E)

Reduction of heme s)-nthesis lnhibition of the phosphatidyl lnositol cycle Blockage of reticulocyte DNA rynthesis Inhibition of B-globin gene expression

Considering the patient described in the previous questlons, treatment with which of the following vitamlns
may be successful in controlling this disorder?

Routine newborn screening identified a child with elevated levels of cr-ketoacids of the branched-chain amlno acids. A certain subset of such children will respond well to which of the following vitamln supplementatlon?

(A) B, (B) B, (C) B, (D) BU (E) B,,

(A) Niacin (B) Riboflavin (C) B,, (D) Bu (E) Thiamine

',$

Another routine newborn screening identified a child

with elevated levels of the branched-chaln amino acids and their o-ketoacid derivatives. In addition, the child
exhibited Iactic acidosis. Which en4.,rne listed below would you expect to be negatively affected (reduced actir,,rty) by this disorder?
also

13-year-old boy is admitted to the hospital due to flank and urinary pain. Analysis demonstrates the presence of kidney stones. The stones were composed of calclum oxalate. Family history revealed that the boys father and mother had had similar problems. Oxalate accumulation arises in this patient due to difflculty in metabolizing which of the following? (A) Alanine

(A) cr-ketoglutarate dehydrogenase (B) lsocitrate dehydrogenase (C) Malate dehydrogenase (D) Succinate dehydrogenase (E) Acetyl-CoA carboxyiase

(B) Leucine (C) Lysine (D) Glyoxl,4ate (E) Glycine

An l8-year-o1d boy was brought ro rhe hospiral by his mother due to a sudden onset of flank pain in hls left side, radiating toward his pubic area. His urine was reddish-brown in color, and a urinalysis showed the
presence ol many red blood cells. When his urine was acidified with acetic acld, clusters of flat, hexagonal

A Russian chi1d, 5 years o1d, was brought to the pediatrician for developmental delay. Biood analysis showed elevated 1eve1s of phenylalanine, phenyllactate, and phenylppuvate. The developmental delay, ln this condition, has been hlpothesized to occur due to which of
the following?

transparent crystals were noted.

A radiograph of the

abdomen showed radio-opaque stones in both kidneys. The boy eventually passed a stone whose major component was identified as cystine. A suggestion for treatment is which of the following?

(A) Acidosls due to elevated phenyllactate (B) Lack of ty,rosine, now an essential amino acid (C) lnhibition of hydroxylating enz)rynes due to accumulation of phenylalanine

(A)

Increased ethanol consumpt ion

(D) Lack of large, neutral amino acids in the brain

(B) Restriction of dletary methionine (C) Utilize drugs that acidify the urine (D) Restrict dietary glycine (E) Prescribe diuretics

(E) Inhibition of neuronal glycolysis by phenylpl-mvate

t4 You have an elderly patient with a history of

A l2-year-d.d boy is brought to the pediatrician because of behavioral problems noted by the parents. Upon examination, the physlcian notices brittle and coarse hair, red patches on the skin, 1ong, thin arms and legs

heart

attacks (MIs) and strokes (CVas). Blood work indicates an elevated homocysteine 1eve1, which ls reduced by the patient taking pharmacological doses of p1-r1doxamine. An enz),rne that would benefit from such

Amino Acid lvletabolism and the Urea C1-c1e ,reatment in lowering homocysteine 1evels is which of -he lollowing? -\) Methionine slrlthase B) N5, Nl0 methyiene tetrahydrofolate reductase

L29

t8 A patient

presents with episodes of flushlng, diarrhea abdominal cramping, and wheezing. His blood pressure and pulse rate are normal during these episodes.

C) Cystathionrne D) Cystathionase

B-spLthase

Physical exam is normal except for scattered telangiectasias. In order to diagnose this problem, a24-hurtne

collectlon for which of the following would be most

appropriate?

E) S-adenosyl homocysteine hydrolase

-{ 3-month-old boy of French-Canadian ancestry ts seen b1-the pediatrician for failure to thrive and poor appetire. Physlcal exam denotes hepatomegaly and a ye1lowing of the eyes The boy had been vomiting and had diarrhea, and a distinct cabbagelike odor was apparent.

(A) Vanillylmandelic acid (VMAs)

(B) Catechols (C) Dopamine (D) 5-hydroxlnndoleacetic acid (5-HIAA) (E) Cortisol

This disorder is due to a defect in the metabolism of ri'hich of the follor,ving amino acrds? ,,A) Alanine 18) Tryptophan

i9 A patient taking

tC) Tyrosrne (D) Histrdine (E) Lysine

a drug for depression experlenced a greatly increased heart rate and sweating after eating red wine and gourmet, aged cheese. These symptoms appeared due to an inabiiity to degrade which of the followlng?

\'tr

Smith had been prescribed a drug to treat his

depresslon. One ol the effects of the drug is to maintain elevated levels of a particular neurotransmitter that has been denved from which of the lollowing amino acids?

(A) Tyrosine (B) Tyramrne (C) Serotonrn (D) Glycine

(E) Glutamate
A 6-year-old boy is slightly anemic and is very sensitive to the sun, to the point where his skin blisters instead of healing normally from sunburn. His condition worsened when he was taking rifampin for a

(A) Tryptophan (B) Tyrosine (C) Giutamate (D) Histidine

(E) Glyclne

Methlcillin Reslstant Staph Aureus. The boy most

1ike1y has a defect

in which of the following bi.ochemi-

l7 A patlent

has a "pill ro1ling" tremor, "cogwheel" rlgidity, bradykinesis, speech difficulties, and a shuffling gait. The chemical that is lacking in this s1,-rdrome is a derlvatlve of whlch of the follou,'ing amino acrds?

cal pathways?

(A) Alanine

(B) Serine (C) tr'rosine (D) Tryptophan (E) Phenylalanine

(A) Glycogen rynthesis (B) Fatty acid oxidation (C) DNA repair (D) Tianscription-coupled DNA repair (E) Heme syrthesis

130 I

Chapter 15
and norepinephrine) and serotonin cannot be sl.nthesized

ANSWERS
The answer is B: 5-hydroxytryptophan. The child has
nonclassical phenylketonuria (PKU). Classical PKU is due to a defect in phenylalanine hydroxylase, leading to accu-

as those pathways require tetrahydrobiopterin Gir,rng

5-hydroxytryptophan blpasses the block in serotonin biorynthesis, and wouid have to be a supplement for these

mulation of phenylalanine derivatives. These interfere with amino acid transport into the brain and can lead to cogrutive disorders rf not treated, usua11y, by a low-phenylalanine diet. However, in nonclassical PKU, the required cofactor for the phenylalanlne hydroxylase reactlon, tetrahydrobioptenn, is deficient. Thls will lead to srmilar biochemical s),mptoms (elevation of phenylalanine derivatives), but, rn addition, the catecholamines (dopamine, epinephrine,

children along wrth dihydroqzphenylalanine (DOPA), which is the hydroryiated precursor for catecholamine biosynthesis. Providing tlrosine will not overcome the block ln neurotransmitter biosy,nthesis. Pror,rding phenylalanine just makes the problem worse. Neither melanin
nor alanine wrll blpass the metabolic biock of this disease.

The role of tetrahydrobiopterin, indicatrng its oxldation and subsequent reduction, in the phenylalanine hydroxylase reaction ls shown below

GTP

+
I

Biosynthesis

((

/n\t"
)

NH"

tscHr-cH-cooPhenylalanine

Tetrahydrobiopterin
(BHa)

Phenylalanine hydroxylase

Hzo
NADH + H*

o
Quinonoid dihydrobiopterin
(BHz)

,*@.,,
Tyrosine

The answer is D: Tyrosinase. The chlld has albinism, a lack of plgment in the skin celIs, which is produced by melanocytes. Melanocyte tl.rosinase (a different isoz)rme

The answer is C:

Citrulline.

The child has ornithine rran,

than the neuronal tyrosinase that produces DOPA for

catecholamine biosyrrthesis) is defective rn albinism. The DOPA produced is then used for pigment production. A lack of phenylalanine hydroxylase leads to PKU. A lack of dihydrofolate reductase is most 1ike1y a 1etha1 event as there are no reported cases of a lack of this en4.me. Tetrahydrololate is not required for the conversion of tl.rosine

to

DOPA

in

melanocytes. NADPH oxidase generates

superoxide, which ls not part of this pathway Homogentisic acid is part of the phenylalanine and tyrosine degradation pathways, and is not involved in albinism.

scarbamoylase (OTC) deficiency, and cannot condense carbamoyl phosphate with ornithlne to produce citrulline (see the figure on page 131). The excess carbamoyl phosphate produced leaks into the cytoplasm where it bypasses the regulated enzyme of de novo pyrimidlne production, leading to excess orotic acid. Thus, in an OTC defect, carbamoyl phosphate can be produced, but citrulline cannot. Since citrulline cannot be produced, the later products of the urea cycle (argininosuccinate and arginine) are also produced at lower 1eve1s than normai. which is an indirect effect due ro the inabilitv to

produce citruillne.

Amino Acid Metabolism and the Urea

Mitochondrion

Cycle f 3I

30, +

HrO

Cytosol

.-->Urine
NH, t'
O
1

E
E

Hco; +
I

NH],

Urea C=
\-.,*

NH, --

,NHzi
cH2
I

c =iNl,

-,

2ArP\13;j::[:?' ,oo"

NH

/lo +Pi
I

synthetase I

Arginase

CHz
I

rcpsrr

cH2NH2
I

CHz
I

CHz
I

cH2NH2
I

H-C-NH2
I

HzN-C-O-P-OI oCarbamoyl

oo llll

CHz

H-C-NH2
I

CHz
I

cooH

CHz
I

COOH

H-C-NH2
I

\HC ^,;"'?'?i'l \,)tt

Argininosuccinate

Ornithine
Ornithine

cooH
Ornithine
NHr

@ \/?* \
lyase I
I

COOH

phosphate

Fumarate

C=O
I

t-

cH2I

NH

cH2I

NH

CHz
I

COOH NH ll .-.*', I C - NH:-CH 1'-. "*'! I CHZ CH,- NH t-l

|

CHz
I

cHt
I

cooH

CHz

CHz

H-C-NH2
I

H-C-NHi
I

COOH

COOH

Argininosuccinate synthetase

CHz

H-C-NH2
I

COOH

Citrulline

Citrulline

Argininosuccinate

r--*- -: I H,N-C-H
CHz
I

cooH

ATP

AMP +

PP1

COOH

Aspartate Answer 3: The urea cyc1e. Reactron 2 is defective in the disease descrlbed in this
case

The answer is C: Bypassing carbamoyl phosphate syn' thetase ll (CPS-ll). The rate-limiting step for de novo plrimidine s)'nthesis is carbamoyi phosphate slrlthetase

ii (cps-il),

which produces carbamoyl phosphate in the figure on page 132). In an OTC defithe carbamoyl phosphate produced in the mito.i.n.y, choniria leaks lnto the cytoplasm, leading to orotlc acid sl,rLthesis as the regulated step of the pathway is being blpassed. The elevated ammonia is not a substrate of
cytoplasm (see the

CPS-II, and while glutami.ne is also elevated, and is a substrate of CPS-ll, higher glutamine concentratlons wiil not overcome enzyfiTe inhibition by its allosterlc lnhibltor, UTP Aspartate transcarbamoylase is the reguiated step of plT imidlne bioslT rthesis in many prokaryotic cells, but not in humans. This step ls necessary for pyrimidines to be sgrthesized starting wlth carbamoyl phosphate. CPS-I is a mitochondrial enz)'me not involved in pyrimidine production.

132

Chapter 15
Glutamine

CO,

2ATP

AO#;JO
:P '^ o o
(actrvation) ?=o
scoA

Carbamoyl phosphate V

[r-

Rsoartate

+ +
Orotate

il,-enee
I

o__\_o
-,.'\
I

OH ilI O-C-C-H
I

(Gtycine)

ruUi

I _^ u-\J

-SCoA Y
I

l.r.o, v
UMP UDP

SCoA

C=O
,1,,

Hippuric acid (excreted)

I**
I
fdCMP

CH,
I

----------+ dUDP
I
I

ooCH"CH"CH"-C,,O

/lCx,

@<-urP

Gr^l

etutamine

l''tp

l/*-"Y,
/ CDP
\R/

NHi

i
I

,,,\'
\2
Phenylbutyrate

'o

B-oxidation

)-'

dUMP

o-".:
Phenylacetate

LdIMP
I

5,10-Methylene-FHo

z-..-r+ dCTp<-ldCDp' (or.nI

\ /4irrD-

I*.*,
+

?o
?-*r, cH,
I

CHz

^..o -t.scoo

t4ttp

y'cols

\orP * ,r,

dTDP

An ovemew of pyrlmidlne syrthesrs, indicating the reguiatron that

occurs at the carbamoyl phosphate syrrthetase II step. 1l carbamoyl phosphate can be generated outslde of thls pathway (as in an ornlthtne transcarbamoylase deficiency), then pyrimidlne slnthesis r,v111 blpass its regulated step, and an overproduction of pl,rlmidines

-J H-3-r.rHr.

Jr,

,Ct' ool

f-t
I

HSCoA

would resuk.

(Glutamine) +

5,:

'

The answer
there is
a

is C: Arginine and benzoate. Whenever

o
t-

CH,

Oi il: -c-NH-C-CH2CH2 -c-NH2 to

ilH

.ct oo
Phenylacetylglutamine

i
i
;

urea cycle defect, arginine becomes an essential amino acid (as its route of sy,nthesis is the urea cycle). Benzoate, along with phenylbutyrate, is given to parienrs

- .. ("*r'""t"9

nirrogen carmolecule (benzoate conjugates with glycine while ryang phenylbutyrate, after activation to phenylacetate, conJugates with glutamine), which is rhen excreted. The reactlons of benzoate and phenylbutyrate with nitrogen containing amino acids are shown above. The excretion
a

with urea cycle defects to conjugate with

Removal of nitrogen using benzoic acid (panel butyraie (panel n)

A) and

phenyl-

.rC-, The answer is B: Reduction

of heme synthesis. The boy is suffering from lead poisoning, which he obtained from eating the flaking paint chips. Lead inhibits the
6-aminolel.r-rlinic acid dehydratase step of heme syethe-

of glycyl-benzoare reduces the glycine levels of the body,

forcing more glyclne to be produced and pror.rding an alternative pathway for nitrogen disposal in the
absence of a lunctional urea cyc1e. Giving lysine or g1u-

sis, leading to reduced heme levels (see the figure on page

133). In addition, the ferrochelarase srep (in which iron

is inserted inro the newly sy,r-rthesized heme ring) is also

tamine
patient.

wiil not help to reduce ammonia

levels

in

the

inhibited by lead. The reduced heme levels reduce the amount of functional hemoglobin synthesized, leading

Amino Acld Metabolism and the Urea

Cycle 133

to the mlcrocytic anemia observed in the chl1d. Lead does not interfere with iron transport or inhibit part of
the phosphatidyl inositol cycie (1lthlum is the metal that does that). DNA syrrthesis is not impaired by 1ead, nor does lead inhibit gene expression of the globln chains. Cytochrome syrrthesis is also decreased and may contribute to the lethargy observed ln the child.

which requires the same five cofactors as do pl,uvate and cr-ketoglutarate dehydrogenase; thiamine, NAD*, FAD, lipoic acid, and coen4/rne A. A subset of patients with this disorder has a mutation ln the El subunit of the errzqe, which has reduced the affinity of the enz).'rne for vitamin B,. lncreasing the concentration of B1 can
therefore overcome the effects of the mutation and a11ow the enz)rme to exhibit sufficient acti\'lty to reduce the buildup of the toxic metabolrtes. While niacin and riboflavin are required for the onzyfire, the mutation in the en4lrne is such that the affinity of these cofactors for the enzyme has not been altered. B,, and Bu are not required for this reaction.

coo-

?oocHz

?*,
cHz

H./ i" ,1,*, \'H

2 6-ALA 6-ALA
dehydratase
I 1

?r, C=O

ll

o=? H:C-H
8

The answer is A: cr-ketoglutarate dehydrogenase. The child has a mutation in the shared E3 subunit of pyruvate dehydrogenase, o-ketoglutarate dehydrogenase, and the branched-chain cr-ketoacid dehydrogenase. AI1 three reactions are oxidative decarboxylatlon reacuons

la, ,rr9
+

cooI

and utllize a three-component enzFne complex, designated as E1, E2, and E3 (see the flgure below). The El subunit binds thiamine pyrophosphate and cataIyzes the decarboxylation reaction. The E2 subunit is a transacylase and is involved in the oxidation-reduction

?,, c-c CCH #, \*/ ril

NHz
H

?oo-

?,, ?,,
il

PorPhobilinogen (a pyrrole)
One of rhe two steps in heme biosyT rthesls that is sensitrve to 1ead.

part of the reaction. The E3 component (dihydrolipoyl dehydrogenase) is shared among all three enzymes, and a mutation in this subunit will affect the actir'rty of all three enzymes. This subunit reduces NAD., using electrons obtained from reduced lipoic acid. The key to solving the probiem is the recognltion that lactic acldosis occurs, which would happen when pyr-uvate
dehydrogenase was defective. None of the other dehydrogenases listed (isocrtrate dehydrogenase, malate dehydrogenase, and succinate dehydrogenase) require

'J"r fhe answer is E: Thiamine. The child has maple syrup urine disease, a defect in the branched-chain cr-keto
acid dehydrogenase step that utilizes all three branchedchain cx-keto acids as substrates. The reaction catalyzed by this er-:z)rme is an oxidative decarboxylation reaction,

the E3 subunit for their activity, nor do they catalyze oxidative decarboxylation reactions. Acetyl-CoA carboxylase caLalyzes a carboxylation reaction, and does not share subunits with the enz)'rnes that catalyze oxidative decarboxylations.

OH
I

RAnswer 8: Mechamsm of q-keto acid dehydrogenase complexes. R represents the portion of the o-keto acld that begins with the B carbon. Three dlfferent subunits are required for the reaction: E1 (o-ketoacid decarboxylase), E2 (transacylase), and E3 (dihydroLpoyi dehydrogenase). TPP refers to the cofactor thiamine pyrophosphate. Lip refers to the cofactor lipoic acid.

Hl
"or,\,, ,r,
i
.-]'-**- r
i

rl-TPP
',-

FAD

(2H)

..

c-Keto
acid

o%J..,

,,

,S. ,,t E3 S-Lip : t' r--.:l-', FAD ... trans Ac G) '..-----' E2

t

Dihydrolipoyl

DH;'.

NAD*

'lc,)

,, {

t\RoH +H+

C=O

H,. 1

.,'
i

o-Keto
TPP

E1 0.i i\
"f

rr"

A"-^- Lip/-SH

SH

coo-

acid DH

l:

"-L.

"",0

HS S-C-R

,,,0. ?

\:,/ /--rR_C_SCoA ,,.::

iCoASH

134 9

Chapter 15

entering the brain (such as tryptophan, ry,rosine, lysine, and leuclne) This alters rhe abiliry of the brain to synthesize proteins, and leads to neurologi_ cal problems Providlng large amounts of these iarfe,
neutral amino acids prevents saturation of the system by phenylalanine, and can be used as a rreatment, along with resrricred phenylalanine diet, for children wlth this disorder. (See J Inhent Metab Dis. 2006 Dec,29(6):732-738.) The developmenral delay does not appear to be due to acidosis, lack of ryrosine, an inhibirion of hydroxylaring enzymes, or inhibition of
neuronal glycolysis.

The ansurer is D: lack of large, neutral amino acids in the brain. The child has pKU. The elevated pheny_ lalanlne 1eve1s in the blood are saturaring rhe 1arge, neutrai amino acid transport protein in the nervous system (L-system), preventing other substrates from

is a diagnostic marker for pKU, but it is not relevant for homocysteine producrron or degradation. Fibrillin
is mutated in Marfan sl.ndrome, but this disorder is not

Marfan slndrome.

fl

The answer is D: Br. Cysrarhionine B-s1,r-rthase is a B, requiring enzyme lthe reaction rs a B-ellmrnation o? the serine hydroxyl group, foilowecl by a B-addition of homocysteine to serine; both tlpes of reactions require the participation of Bu). In some murarions, the affinity of the cofactor for the errzqe has been reduced, so

significantly lncreasing the concentration of the cofactor

wrll a1low the reaction to proceed. The enzyme does not

require the assistance of B, , B, B, (niacin), or Iyze the reaction.

B, to cata_

l0

1Z

The answer is E: Homocystine. The boy is exhibiting

the symptoms of homocystlnuda, usually caused by

defect

in

condense homocystelne with serine to form cystathionine. An inability to catalyze thls reaction will lead to an accumulation of homocysteine, whlch will oxidlze to form homocystine. The elevated serine can be metabolized back into the glycoI1,tic pathway Methion_ ine wrl1 not increase in blood as the homocysteine pro_ duced is converted into homocystine. phenylpyruvate B-synthase

will

cystathionine B-sprthase. Cystathioninl

The answer is D: Glyoxylate. The boy has primary oxaluria type I, an autosomal recessive trait, which is a defect in a transaminase that converts glyoxylate to glycine. If thls transamlnase is defective, glyoxylate wr11 accumulate. The glyoxylare wrll then be oxidized to oxalate, which, in the presence of calcium, will prectpr_ tate and form srones in the kidney The metabolic parh_ way for glycine being converted to glyoxylate is shown be1ow, and the enz)rme that catalyzes thrs reaction is
the D-amino acid oxidase. Alanine, leucine, and lysine metabolism do not give rise to oxalate.

-o g-6r"t -5;;'/ llto-g OH NH;

Threonine

CH.

glutarate
Answer 12

o_Hydroxy_

B-ketoadipate

Amino Acid Metabolism and the Urea

eYani

Cycle I35

rnllir:
-s

t3

The answer

is B: Restriction of dietary

methionine.

no:

The boy has cystlnuria, elevated levels of cysrine in rhe

urine, due to a defect

in a kidney

transporter that

:JI)

removes cystine from the urine and sends it back into the blood. Due to this, the concentration of cystine in the urine is higher than normal and reaches levels
o,

but will not lead to significantly elevated homocysretne S-adenosyl homocysteine hydrolase is the enzyme that converts S-adenosyl homocysteine to homocysteine and adenosine; lack of its activity will lead ro a reduction, not an increase, in homocysteine leve1s.

ln
c

Oil rr,

llra

---;ItLtLld. sc ia:..1 s no:

11ry.

::t i:
LC Lt'

close ro its solubility hmir. Cysteine is derived from methionine, so a reduction in methionlne levels will reduce cysteine levels, which then leads to a reduction in cystine levels. Increasing ethanol content will lead to dehydratlon, which will increase the concentration of cystine in the urlne, leading to increased precipitation. This would also be the case if the urine were acldified (acidification also reduces the solubillty of the cystine stones). Restricting glycine is not effective, as glycine is not a precursor of cysteine biosynthesis. Prescribing diuretics would force the boy to urinare more frequently, and would raise a risk for dehydration, which would lead ro possible elevarion of cystine concentrations.
t+:

t5

Tyrosine. The boy has the inherited disorder tyrosinemia rype I, which is a defect in fumaryIacetoacetate hydrolase, the last step in the degradation pathway for tyroslne (see the figure below). In irs acure form, this dlsorder will lead ro liver failure and death within 1 year of life. The accumulation of inrermediares in the tyroslne degradation parhway triggers apoprosis
The answer is C:

'f'. ( ( ) Ycu,-cH-coo\ \_-,/ / D-C

Phenylalanine PKU
phenytalanine hydroxytase

/7T\
+

-.t --

"r1-

a:D1-

The answer is C: Cystathionine B-synthase. Cysrathionine B-synthase has a requiremenr of pyridoxal

phosphate, and tn about 50% of the cases of defective synthase enzymes, increasing the concentration of B. can overcome the effects of the mutation on the..rrylTI.I While a defect in methionine synrhase will lead to elevated homocysteine (see the figure below; cystathionine B-synthase is enzyme 3 and methionine sl,nthase is errzpe 1), thls enzyme requires B,,, not Bu. A defect in N5, N10 methylene terrahydrofolate reducrase will also lead to elevated homocysteine, but that enzyme has a requirement for NADH, not vitamin Bu. A defect in cystathionase (another Bu requiring enzyme) will block the degradation of cystathionine, which will accumulate,

)a-ih-

NH"

J\\Tl
):1
iS

H2-CH-COOTyrosine
Tyrosinemia

t"

-s1na

ll Il

i
HO

prp

ryrosine aminotransferase

cH2-c -coopHydroxyphenylpyruvate
I

il

[.-r"o^
N5,N1o-methylene-FH4\,

FH, PP' P' ' C/-+laetnionine ^lt l nethvlFH^48,, + + Dimethyl gtycine ( af, ./ ' Betaine HomJcYsteine' )'a \R-CU.. ,> ->-^,/ a" ,/ s-adenosyt himocysteine @/ aa.,lo""ne t -'-'(-"""n"
f

ctycine }- Serine

i
r

-oH

HO2-C
Alcaptonuria

fL \\__/,/
II
I

-cH2-cooHomogentisate

Homooentisate oxidase

+ +
Tyrosinemia I

c+

Cystathionine

frmaryracetoacetate hydrotase
I

'li

;g*5"#

"6fa-Ketobutyrate.

NH.

rcOC

CH= CH-COOFumarate

"\g

cH3

- c-cH2- cooAcetoacetate

f36

Chapter 15
of the hepatocytes, leading to complete liver failure. The

or

phenylalanine. The figure below indicates the

yellowing of the eyes (jaundice, due to accumulated bilirubin) ls a result of liver failure. None of the other amino acids listed (alanine, tryptophan, histidine, and lysine) contribute to the formation of intermediates of
the phenylalanine and tyrosine degradative pathways.

biosynthetic pathway of DOPA and the catecholamines.

Phenylalanine

on"
hydroxytase

f-

auo

I*rr,
HoTz)

ffi

ffr" answer is A: Tryptophan. Most drugs used ro rrear depression do so by elevating serotonin levels, and serotonin is derived from tr;.ptophan (see the figure below). Tposine is the precursor fot catecholamines,
whl1e glutamate is the precursor of GABA. Histidine is the precursor for histamine, while glycine irself acrs as a neurotransmitter in the brain.

\,,'^cHl
tYrosine nyoroxvrase

t" CH O_ tc'
o
il

NHI

l-Tyrosine

||:+

BH+

AU,

HoYz\
<-\--lcH2-cH-tiH3

;,dH:

<-\-/
H

coo-

---->or

Nicotinamidemoietv
NAD(P)

I ll cH o-\ \-, cH/ i'"c'.. ' ll i

:*o-i

T't

Tryptophan

o, --,[r- auo I H2Off.>BH2

tryptophan hydroxylase

oopa decarboxylase

Dopa

lprp
I

'io-,

r "-r/ -4.)r-=rcH2_cH_NH3

OH Neurons
-.1

l-"ia", Y --'

(/---,l,l
H

HO

i6q=

cHi

cH" -'ruu5

5-Hydroxytryptophan

ele [-> CO,

I-:

Dopamine

i '--'

DoPA decarboxytase

'"G-JcH2-cH2-fiH3
H

Serotonin

Norepinephrine The answer is C: Tyrosine. This patient has Parkinson disease, which is a problem with dopamine slmthesis in the substantia nigra. Dopamine is derived from

tl,rosine. Tieatment with DOPA in the initial stages of the disease provides relief from the qrmptoms. DOPA cannot be slmthesized from alanine, serine, try,ptophan,

OH

'"h

\,^?*'/
OH

cH, ici; -\tdH.'----:-

Epinephrine

Amino Acid Metabolism and the Urea

Cvcle 137

:es

the

lmines.

The answer is D: 5-hydroxyindoleacetic acid (5-HIAA). This patient has the classic presentation of a carci-

noid tumor. This tlpe of tumor secretes serotonin that causes these classic symptoms. The breakdown product of serotonin is 5-hydroxyrndoleacetic acid (5-HIAA, see the flgure below). Elevated levels of 5-HIAA in the urlne confirms a hlgh level of serotonin and the diagnosls of a carcinoid. VMA and/or catechols would be elevated if the patient had a pheochromocytoma

producing epinephrine or norepinephrine (rhe t.i.!--. are degradation products of these neurotransmi'.-.:,r also seen in the figure below). The symptoms do noi match a pheochromocyroma, particularly due to the lack of increase in heart rate or blood pressure. Dopamine is depleted in Parkinson disease, not in this condition. Cortisol levels would be high in Cushrng syrrdrome, but not under these conditions.

,/t--nu

\r"\N,/

""'-i;;-" x--If
H

Ho)-- oH ,/r'r \ I + ,o19cH-cH2-frH3

Tryptophan

I g"g<-f>BH"

-+

o, ---,lr- eH, -

tryptophan hydroxytase

M^o/ /*,,
Ho>-----1

Norepinephrine

sar\corur

""G--rcHz-cH-fiHs
H

:;,\
o
.

*o{O}3*-di,
\--l
lor,o",,on

oH

cH3o.

5-Hydroxytryptophan
I

,o{O } \-7
I

nu vtt

3',- cr,-

*r,

f'+69, DOPA decarboxytase i 'oYZ)I--tt-cH2-cH2-r{H3 MAo_A

ere
,drenal

Ho)- I
o

NHI<-1

I MAO

\.\_/

tedulla

s",Jonin

,"GlcH,-B-H
5-hydroxyindoleacetaldehyde

---}.*,.

,o1O!3r-"oo-

oH

\\-7

Oxidation

""ry^'
3-Methoxy-4-hydroxymandelic acid (Vanillylmandelic acid, VMA)

+o
H

[,-

Nrao.

l.+ruaoH

5-hydroxyindole acetic acid (5-HIAA)

Panel A shows the generation ol 5-HIAA from serotomn degradation, while panel B indicates the generatlon of VMA from catecholamine
degradation.

The answer is B: Tyramine. Tyramine is a degradation

product of tyrosine (decarboxfated tyrosine), which, when elevated, will lead to norepinephrine release. Tyramine is found in red wine and aged foods such as certaln cheeses. When lngested, tyramine rs degraded by monoamine oxidase to a harmless compound,

However,

and excessive norepinephrine release does not occur. if a patient ls taking a monoamine oxidase

inhibitor (MAOD, it is possible that ryramine does nor get degraded approprlately. MAOIs which covaientll' modify (as opposed to being comperirive inhibirors) the enz),rne are very useful medications for atypical

138

Chapter 15 depression that is unresponsive to other modalities. Unfortunately, MAOIs have multrple interactions with many other medicatlons and foods. A high tyramine level leads to a greatly elevated blood pressure due to the release of norepinephrine. Patients on MAOIs need to avoid foods high in tyramine, such as cheeses (aged and processed), red wine, caviar, brewer's yeast, miso soup, dried herring, and aged meats. MAOIs have no effect on glycoproteins or cholesterol.
heme precursors in skin cel1s that are easily converted

to radical form by the energy in sunlight, and which severely damage the ce1I. The drug the boy is taking
is metabolized via a cytochrome P450 system, which is induced when the drug first enters the circulationInduction of P450 systems induces the synthesls of heme, ieading to increased concentrations of the heme intermedi.ates and an increased sensitlvity to the effects of these intermediates as induced by sun1ight. The anemia is due to reduced heme levels in the red blood cells. This disorder is not due to defects in DNA repair, glycogen metabolism, or fatty acid
metabolism.

synthesis. The boy has porphyria, a reduced ability to synthesize heme The supersensitivity to the sun is due to the presence of
The answer is E: Heme

Chapter 16

Phospholipid
This chapter quiTTes the reader on the biological roles of phospholipids, sphingolipids, and glycosaminoglycans. Diseases relating to these large molecules willbe the focus of this chapter
(A) Dipalmitoyl phosphatidylcholine

(B) (C) (D) (E)

Palmitate containing ceramide Sphingosine

Sphingomyelin Diacylglycerol

QUESTIONS
Select the single best answer. ;.1.1., A patient presents with rapidly progressive weakness of the lower extremitles, Ioss of deep tendon reflexes, respi-

Considering the case in the previous question, the malor function o[ the suspension utilized to improve breath-

''

ing is whrch of the followlng? (A) To allow oxygen exchange with red blood cells

ratory distress, and autonomic dysfunction following a flulike illness. This disease is an autoimmune inflammatory reaction to tissue made up chiefly of which of the

(B) To facilitate carbon dioxide extraction from red

blood
ce11s

followlng chemical structures?

(A) (B) (C) (D) (E)

.:Z;,. The

High-density lipoproteins
Elastin

(C) To reduce surface tension at the air-water interface (D) To stabilize the structure of lung cells (E) To facilitate blood flow through the lung
A 9-month-o1d child is taken to the pediatrician for lethargy and poor feeding. The physician notes a cherry-red spot in the childs retina. The baby seemed fine for the first three to six months, then began to have problems swallowing, overreacted to loud sounds, seemed to have problems with her vision, and began losing muscle mass and strength. Measurements of which two metabolites is critical to correctly diagnose this disorder?

Sphingolipids Glycoproteins
Glycogen

above patient is in the recovery phase of her i11ness. She wants to "natura11y" help her body recover using dietary methods. Which of the following foods is best in providing the chemicals needed to regrow the affected
tlssues?

(A) Soybeans (B) Calves' liver (C) Pork kldney (D) Green leafy vegetables (E) Potatoes
3l

(A) (B) (C) (D) (E)

GM2 and globoside GM2 and GM3 GMl and globoside GMI and GM2 Globoside and sphingomyelin

A newborn infant had trouble breathrng at blrth.

The

'.:f.t,

infant was 3 months premature. The physicians [reated the lnfant with a soiutlon, which was directly injected into the lungs. Within seconds, the infant responded with much improved breathing. A major component of this solution is which one of the following?

Considering the child described in the previous question, a diagnosis of Sandhoff disease was made. This results ln a loss of which of the following enzl.rnatic
activities?

(A) (B) (C) (D) (E)

Hexosaminidase A and Hexosaminidase C Hexosaminidase B and Hexosaminldase C Hexosaminidase A and Hexosaminidase B Hexosaminidase A and sphingomyelinase Hexosamlnidase B and sphingomyelinase

139

L4O 7

Chapter 16

Consrdering the child described in the last two qucstions, multiple enzymalic activilies are lost. This rs due to

which of the following? (A) A common operon for the two genes conlalns a mulaLion in the promoler rcgion (B) An inactivating mutalion in an activator for the lost
enz) mJl i( acti\ ilies

(A) Galactosylceramide (B) Sullatide (C) Glucosylceramide (D) Sphingoml,elin (E) Ceramide
10

(C) A transcriptional actl\.ator is inactivated (D) A common mutated subunit is present in the multiple actirities (E) A transcriptional repressor is activated

The sphingolipidoses, as a class, are most similar to r,r,hich one ol the following disorders?

A 4-month-o1d infant is brought to rhe pediatrician for a variety of problems. The chrld is frequently irritable, smal1 for age, r.omits frequentll', and displays hlpotonla, as well as hlperesthesia (audrtory tactile, and r..isr-ral). Liver and spleen size are nonr-1al. As the chtld ages, his condition worsens, with rapid psychomotor deterioration, seizures, and bltndness. This disorder is caused by an accumuladon of which ol the follou'ing rn neuronal lysosomes?

(A) Gluco se- 6 -phosphate de\dro (B) von Gierke disease (C) Zelhveger s1'ndrome (D) MELAS (E) 1-ce11 disease
11

genase defi cienc,v

A chrld has been diagnosed with Ta1'-Sachs disease , Lr which a particular liprd accumulates u'ithin the lysosomes. The component ol this lipid which cannot bt removed in the lysosome is which of the fol1ow-ing?

(A) Galactosylceramide (B) Sulfatlde (C) Glucosylceramrde rD) Sphingomyelrn (E) Ceramide

(A) Ceramtde (B) Sphingosine (C) Fatty acid (D) Glucose I Er N-acer) Igalactosrmine
12.

A 6-month-o1d boy ls brought to the pediatriclan due to a large stomach. The doctor noticed splenon'regaly,
with no pain. The boy was always trred and had anemia. The boy also has thrombocytopenia and brulses easily X-rays show a deformity of the distal femur, as shown below. Thls disorder rs caused b)'an accumulation of which of the followrng in macrophage lysosomes?

A depressed patient is prescribed lithium by his ps1'chlatnst. The eflect of lithium is to block the generetlon
of u,hich of the followi.ng?

(A) Diacylglycerol (B) inositol trisphosphate (C) Inositol bisphosphate (D) lnositol phosphate (E) Inositol t3
An alcoholic patient with advanced cirrhosls presents with spur cell anemia. For virtually all cell types and
organeiles, the phospholipid compositlon of the inner and outer leaflets of the membrane is different. The spur cell anemia is the result of the loss of one potential beneflt of such phospholiprd asltnmelry. Which of the follou,ing best explains thrs benefit? (A) To vary the melting temperature of the membrane

(B) To represent all phosphohpids

membrane

specres

within the

(C) (D) (E)

To mark ce1ls for recognition by outside systems To distrnguish between intracellular organelles To prevent lusion of intracellular organelles

l4

Phosphatidylinositol contribules to phospholipid bilayer asynmetry by being in the inner leaflet of membranes, facing the cytoplasm of the ce11. This is most likely due to whlch of the following?

-:- -.-. : --:: -

^--

: C

'-...--- :14 ....':-;: c\'-<:. : I::os.tol is r-en similar 1n struciure to giucose and co,rld compete \\irh glucose for bindrng of ligands io ihe extraceliular surface Phospharidl-linositol acts as a substrate for intracel_
1ular processes

-::_--. -:. - - .: -.-_-..-

.---::1 :-: l.l:--..:--,-.:-- l+l

t8
-{ 2i-r-ear-oid \\ oman sees her phr.sicran ciua :!- ,.,.:-la -::: and tiredness. She has rrngling and numbness ,: ::: fingers and roes, loss of balance and falhng. ;rnd .b,-.:
.

D', Phosphatidylinositol binds ro phosphatidylserine, anorher inner ieaflet specific phospholiprd E', Inosrtol inreracts with intracellular actln, linking the inner leaflet to a cel1,s cytoskeleton

lowing?
(B

r.-iston. sometimes dorrble r.rsion. Her ophthalmolc_ _.. has diagnosed opric neurlrrs in her An tviru of rhe b::. shows "skip 1esrons." The component that is prinL;.:--. defectir.,e in this patienr is composed olwhrch o1 ,h. -.
_

t5

The use ol proteoglycans

in

s1,novia1

adi-anrageous due to the ability of the proteogiycans ro l-orm w.hich type of interactions with other components of rhe fluid?
:! -

fluid of .;oints is

(A) Phosphotrpids and proreins r Triac) lgl; cerol and prorein (C) Phospholipids and triacylglycerol (Dt Crn*,,orides and prorein (E) Triacylglycerol and gangliosides
19

(A) Drsulfide and ionic bonds (B) Hydrogen and ronrc bonds (C) Covalent and ionic bonds (D) Covalent and hydrogen bonds (E) Disulfide and hydrogen bonds

t6

ns ps\

reration

Your 52-year-o1d male patienr, an avid soccer player in his youth, who had severai knee injuries, has been com_ plaining of knee pain for the past 6 months. The knees are tender, stiff, and feel warm when touched. He wants long-term rehef, not just short-term relief you suggest thar rhe parienr take which of the following to try and reduce the knee pain, for the long term?

does nor have sysremic lupus erythematoiis (SLE Examination of the proteins in her bloocl should finc antlbodies directed against which of the follou.ing? (A) Cytochrome c (B) Phosphollpids (C) DNA

A woman has a history of premature mrscarriages (ihree thrombocytopenia, and several episodes of leep ,e,t thrombosis. She has a positlve lupus anticoagulant bu:

(D) nNe splicing proreins

(E) Ribosomes
20

(A) (B) (C) (D)

Asplrln
Acetamlnophen

Sphingomyelln
Glucosamine

(E) Inosltol

l7
resents es and

: inner
re
r1

Children with eirher l-cell disease or Hurler syndrome show very similar cltnical features. One method to dis_ tinguish berween rhe rwo is to f,nd which of the fol1owing eievared in the blood?

An athlete presents to the ER wtth sudden pain in hrs calf-after hearlng a "popping noise,,, and tnability to push off wlth his toes when he tries to run. He gives a history of having a "corrisol shot,, in his heel area for Achilles rendonitis and he just finishecl a course of ciprofloxacin for chronrc prostatitis. physical exam reveals a mass in the superior posterior lower 1eg and an inability ro planrar flex hls foot. Biopsy of the Achil_ 1es tendon would be expected to ..u.ri-fib.otic areas, neorrascularization, and an increase of which of the fol_ lowing in the extracellular matnx?

spur
ben-

he fo1-

)Iane

(A) Heparan sulfate (B) Short-chain dicarboxyhc (C) Lysosomal hydrolases (D) Dermatan sulfate

acids

in the
InS
S

(E) Cytochrome c

(A) Cholesterol (B) Clycosaminoglycans (C) Triglyceride (D) Sphlngosine (E) High Density Llpoproteln (HDL)

ilayer
:anes,

v due

142

Chapter 16

ANSWERS
.

the tlpe I1 ce1ls within the lung have not yer begun ,mthesizin7 surfactant, so the application of surfactant to the baby will aiiolr, this compound to be present until the type II cells begin their s).rithesis of thls complex.

The answer is C: Sphingolipids. This patient has the classic symptoms of Guillain-Barrd syndrome which is an inflammatory autoimmune neuritis wherein T-ce1ls formed in response to a viral illness mistakenly attack

The

the myeiin sheath of perlpheral nerves. The myelin sheaths are composed primarily of sphingolipids and phospholipids and do not contain high-density lipoproteins, elastin, glycogen, or
a

significant leve1 ofgly-

ma.1or phospholipid in surfactant is dipalmitoll phosphatidylcholine (DPPC), and rt is complexed utt: a number of small protelns (surfactant proteins A, E. and C). While small amounts of sphingomyelin ma1'b. present in surfactant, DPPC is the major componenThe sfructure of DPPC is shown be1ow.

coprotein. A view of demyelination is shown below.

o
ll

3 H2c-o-c-(cH2)14-cH3
I

cH3-(cH2)14-c-o-cH
H2C

-O - P- O-

CH2

CH2

- N-

fr. CH3
cHs

the major component of lung surfactant

Nerve fiber

P\

ry
rl \

The answer is C: To reduce surface tension at the Myelin Nerve fiber

air-water
of surfac-

interface. The phosphollpid-protein mixture

tant interacts at the surface of lung cells, allowing expanslon and contraction due to reduclng surface tension a, the air-water interface (see the figure below). Surfactan: does not affect oxygen exchange with the red blood ce1ls nor does it a1low carbon dioxide removal from such cel1s Surfactant does not stabilize lung cell structure (although

lt is essential for the function of the lung ce11), nor it facilitate blood flow through the lung.

does

yJ
Without lung surfactant,
sac collapses. Ten times the normal pressure is needed for reinflation.

it,-

'1

lrnl"t"o
+

t"rrrJ.*-'l

,/

'!*-/

An overview of demyelination. Panels A and B depict normal conditions, whereas panels C and D show the slow disintegration of myelin, resuiting in demyelination and a loss of axonal functlon.

The answer

is A: Soybeans. Foods considered the

highest sources of sphingolipids include dairy and soy products. Foods highest in phospholiplds include those high in leclthin, such as eggs, soy, and wheat. Sphingolipids and phospholipids are found mostly in neural tissue. Other organs and muscle do not contain as high a quantity of these lipids as do neural tissues.

Lung surlactant reduces the surface tension of water (fluid) Iining the surface of the aveolar sac, preventing collapse.

JL

Less pressure is needed

to reinllate sac when

surfactant is present.

;ti

The effect of lung surfactant

The answer is A: Dipalmitoyl phosphatidylcholine. The patient was treated with an artificial preparation of iung

surfactant, which reduces surface tension within the lung at the air-water interface. In premature newborns,

The answer is A: GM2 and globoside. The cherryred spot ls indicatlve of either Tay-Sachs disease (an autosomal recessive disorder ieading to a loss of hexosaminidase A lhex A] activity) or Sandhoff disease (an autosomal recessive disorder leading to a loss of both hexosaminidase A and B lhex B] activity). With just a

..:,:,,!

PhospholipidMetabohsm 143
loss of hex A activity, GM2 would accumulate. With a loss of hex B activity, globoside and GM2 would accumulate. Thus, by measuring the leve1s of GM2 and globoside, one can distinguish between Tay-Sachs and Sandhoff disease. A loss of either hex A or hex B would not affect GM1 or GM3 degradation.

C.

The answel is

(:

Hexosaminidase A and Hexosaminidase

B.

In Sandhoff disease, both hex A and hex B ac::-. -ties are 1ost. The mutatlon in Sandhoff disease does n.-: affect sphingomyelinase activity, and there is no her..-

saminidase C activitlz Sandhoff di.sease is one of man-. which affect sphingollpid metabolism. An ovenieu .-r the sphingoiipldoses is shown in the flgure belour

disease
e?

.-_

,:

Mtachromatic leukdystrophy

1S

ru-acetyl galactosamine

Ceramide
Phosphoryl

lTl
^^^

srrat"
Enzymatic action

Gahctose

@ etr*"*

C N-acetyl neuraminic acid

@
choline

XXX Metabolic block

Answer 6: A summary ol the sphingolipidoses in diagrammatic lorm

tffi
F.,"!:ii!4

144

Chapter 16

"7,' The answer is D: A common mutated subunit is present gene

in the multiple activities. The hexosaminldase A encodes the hex A protein, and the hexosaminidase activlty requires a complex of hex A and hex B proteins;
hexosamlnidase B actrvity only requires a complex ol hex B proteins. Tay-$26[t disease is a defect in the hex A protein, affecting only hex A actirdl)r Sandhoff disease is a defect in the hex B protein, which alfects both hex A and hex B actit'ity, due to the sharing of a common subunit belween the two proteins. hex A and hex B are no[ in an operon (which is only operatlve in bacteria); rn fact, the genes are on different chromosomes. There ls an activating proteln for hex A actil'.ity, but not hex B actil' ity (a loss of the activating protein is known as Sandhofl aclivator disease, with s1'mptoms very similar to TaySachs dlsease). There are no mutatjons in iranscriptional control proteins (either an actlvator or inhibltor) in TaySachs or Sandholf disease. The interactions ol the hex A ancl hex B proteins are shown in the f,gure below'
Sandhoff activator Protein

B gene encodes the hex B protein. Hexosaminidase A

in degradrng ceramide leads to Farber disease, a defect In ceramidase. Farber disease is simrlar to Krabbe disease. but often presents with hepatomegaly and splenomeerl' ' See Tabie 16.1 in the next answer for a summanthe sphingolipidoses and the material that accumui"'' within the lysosomes. Additionally, the figure associat'wrth the answer to questlon 6 of this chapter depicts t: metabolic blocks ol the sphingolipidoses.
The answer is C:

"'

Glucosylceramide. The child has

lo:

'

+
Block in Sandhoff Oisease

lo
Block in Tay-Sachs disease

which removes glucose from glucosylceramide T ' accumulation of glucosylcerebroside in the lysoson-" leads to the observed symptoms. Defects in degr:': ing galactosylceramide lead to Krabbe disease, rvh-d.oes not result in hepatomegaly and splenomegah defect in degradlng sulfatide leads to metachrom;' leukodystrophy, which has different symptoms tll'' what the child is experiencing. A defect in the degr*-. tion o[ sphingomyelin leads to Niemann-Pick dise:'" with a different set ol symptoms than that seen u-'. Gaucher disease. A defect in degrading ceramide 1e:'- ' ro Farber disease. a defect in ceramidase, with n'"' severe symptoms than those observed in Gaucher c 'ease. Table 16-1 summarizes the sphingolipidoses '

of Gaucher disease, which is a defect in a glucosid--''

hexosaminidase A (oz 0z)

* f
+
I

GMz

Ceramide-Glc-Gal-NAcGal
Sialic acid

enzyme defect, and the material that accumulates L ' lize rhis rabie with the figure associated with the &rS" :. to question 6 of this chapter lor a thorough understa:': ing o[ the consequences of the sphingolipidoses. 10 The answer is E: l.cell disease. The sphingolipido"' and l-cell disease are both lysosomal storage diseas"

Hexosaminiciase A or B (Ba)

Block in Sandhofi disease

whereas the other disorders listed

do not

rnv"'

lysosomal dysfunction. Mitochondrial myopa'encephalopathy, lactic acrdosis, and stroke (MELASI -' mitochondrial drsorder, and Zellwegers i.s a disorder

Globoside ceramide-Glc-Gal-Gal-NAcGal
Subsrrate specificities of hexosaminldase A and B, and the functlon of rhe actir.'ator protein. Defects in the B-subunlt lnactlvate both hex A and hex B actirtties, leading Lo GM2 and globoside accumulalion' A defect in Sandhoff activator protein also leads to GM2 accumulalion, as hex A acrir''ity is reduced. Defects in the cr-subunii only inactivate hex A actir.rty, such that hex B activity toward globoside ls unaffected'
G1c, glucose; Gal, galactose; NAcGal, N-acetylgalactosamrne

peroxisomal biogenesis. G6PDH (glucose-6-phosph''' dehydrogenase) def,ciency and von Gierke disease ::' si.ngle gene mutatlons which do not alter lysosol' function (although type 1I glycogen storage dise:-'
Pompe dlsease, is a lysosomal storage disease).

l:l

', '.

The answer is A: Galactosylceramide. The child has Krabbe disease, a mutation in a galactosidase, which cannot remove galactose from galactose cerebroside (an inability to break the bond between galactose and ceramide). The buildup of galactose-ceramide leads to
the neuronal and muscle damage seen in the child An inability to degrade a sulfatide would lead to metachromatic leukodystropy, which has very different s)nnptoms than Krabbe disease. An inability to degrade glucosylceramide leads to Gaucher disease, again, with a very dlfferent disease progression than that seen with Krabbe disease. A defect ln the degradation of sphingomyelin leads to Niemann-Pick disease, with a different set of
s),'rnptoms than that seen w1th Krabbe dlsease A defect

fhe answer is E: Irl-acetylgalactosamine. Tay-Sach- ' a defect in hexosaminldase A, which removes the :' minal N-acetylgalactosamine residue from gangho--'' GM2, producing the free sugar and GM3. Hexosam-: ceramide, sphingosldase A does not cleave glucose, or the fatty acyl component of ceramide from GM2: ' speci fi c lor N-acetylgalactosaml ne

1,2

The answer is E: lnositol. Lithium primarily inhi: -' the phosphatase which converts inositol phosphate free inositol, thereby disrupting the phosphatrdyllnos- ' cyc1e, leading to increased levels of the intermediate' ' the cycle, whlch are often signali.ng molecules. Lithi" does not allect the generation of diacylglycerol, in;to1 tnsphosphate (IPr), inositol bisphosphate (lP.r

PhospholipidMetaboiism
Table 16-1. Defective enzymes in the gangliosidoses

1+;

Disease =-rcosidosis
3eneralized gangliosidosis ay-Sachs disease ay-Sachs variant or Sandhoff

Enzyme DeficiencY
u-Fucosidase
G

Accumulated Lipid
Cer-Glc-Gal-Gal NAc-Gal : Fuc H-isoantigen
Cer-G lc-Gal(NeuAc)-Gal NAc:Gal
G

n,,-B-ga

lactosidase

M 1 ganglioside

Hexosaminidase A Hexosaminidase

Cer-Glc-Gal(NeuAc):GalNAc GM2 ganglioside Cer-Glc-Gal-Gal:GalNAc Globoside plus GM2 ganglioside

Cer-G lc-Gal :Gal

risease :abry disease

AandB
u-Galactosidase Ceramide lactosidase
(B-galactosidase)
G

lobotriaosylceramide

leramide lactoside lipidosis

Vetachromatic leukodystroPhY (rabbe disease
Gaucher disease

Cer-Glc:Gal Ceramide lactoside

Cer-Gal: OSO. 3-Sulfogalactosylceramide

Arylsulfatase A
B-Galactosidase
B-G

Cer:Gal Galactosylceramide
Cer: Glc
G

lucosidase ingomyelinase

lucosylceramide
i i

Niemann-Pick disease
Farber disease

Sph

Cer: P-chol ne Sph

gomyel

Ceramidase

Acyl: sphingosine Ceramide

VeuAc, /V-acetylneuraminic acid, Cer, ceramide; Gic, glucose; Gal, galactose; Fuc, fucose: site of deficient enzyme reaction

inositol phosphate (lP); it affects just the conversion of IP to free inositol and a phosphate.

phosphatidylinositol cycle). As such, it must face the cytoplasm of the ce11 such that when the inositol
phosphate derivatives are produced, such as IP, they can move to their target receptors to elicit a cellular response. lnositol contains six hydroxyl groups and

t3

The answer is

(:

To mark cells for recognition by outside

systems. By havlng different phospholipid compositions of the inner and outer leaflets of membranes, one can utllize phospholipid head groups (which face the aqueous phase of their leaflet) as markers for "inside" and "outside" the membrane structure. For example, the exposure of phosphatidyl serine on the "outside" of red blood cells as is seen ln spur cel1 anemla is a signal for the removal of the cells from circulation by the spleen, as the serine residue should be facing the "inside" of the red blood ce11. Spur cells are large red blood ce1ls covered mth spikelike projections from preferential overexpansion of outer membrane components, leading to a spurlike shape. Movement of the phospholipid rs a signa1 of cell aging. The melting temperature of the membrane is better determined by the fatty acid composltion of the phosphollpids, not the head group composition. Not all phospholipids are represented in all memhranes (for example, cardiolipin is found almost exclusively in the mitochondrta). Assymetric phospholipld compositions do not distinguish one organelle from another (that ls primarily due to protein content), and assymetry in phosphollpid composition may promote fusion (r'esicles need to bud from and fuse with other membranes, particularly in the Golgi apparatus).

is a very hydrophilic molecule. lnositols structure

is

quite different from glucose (there is no carbonyl group in inositol), so it is unlikely that glucose and inositol

would compete for binding to the same receptors. Phosphatidylinositol does not bind to phosphatidylserine in the inner 1eaflet of membranes. lnositol also does not
interact with the actin cytoskeieton.

t3 The answer is B: Hydrogen and ionic bonds.

The

hrgh concentration of negative charges pror'rded by the proteoglycans attracts catlons that create a high
osmotic pressure within cartilage, drawlng water inlo this specialized connective tissue and placing the co1lagen netlvork under tension. The water remains due to hydrogen bond formation with the proteoglycans. At equilibrium, the resulting tension balances the swelling pressure caused by the proteoglycans. Cartilage can thus withstand the compressive load of weight bearing and then re-expand to lt previous dtmensions when that load is relieved. Disulfide bonds and covalent bonds do not play a role i.n proteoglycan stabilization of jornts.

,t+ The answer is C: Phosphatidylinositol acts as a sub'

strate for intracellular processes. Phosphatidylinosltol

is used as the substrate to provide signaling molecules in response to the appropriate stimuli (the

t5 The answer is D: Glucosamine. While aspirin and acetaminophen may provide short-term relief, the use of glucosamine may help to rebuild the proteoglycan layer ln the knees, reduclng the osteoarthritis (although medical studi.es are controverslal concernrng the use of glucosamine and glucosamine

146

Chapter 16

long carbohydrate chalns, which .o*irt of repeat_ ing disaccharide units (see the figure below). Note the

sulfate, in rems of providing relief from joint pain). Sphingomyelin and inositol are not important compo_ nents of the cartilage in joints. proteogiycans contatn

inclusion of glucosamine derivatir.,es in three of the five

repeal ing disaccharide unirs.

t7

The answer is C: Lysosomal hydrolases. In I_ceil disease. the lysosomal hydrolases are mistargeted and are excreted

from celis into the circulatron. As the pH of the blood is above 7 and the pH optlmum ol these enz)rynes is around
5, there is no activity of the hydrolases in blood. in Hurler sl,ndrome, a defect in the degradation of mucopolysac_ charides, there is an accumulation of der-rnatu., u.rd h.pr_ ran suifate in the urine, but not in the blood. Short_chain dicarboxrylic acids are produced wlth a defect in medlum

Glucuronic
acid

B(1

*3)

chain ac14-CoA dehydrogenase, and cytochrome c release ce1ls from mitochondria is the sig_ nal to initlate apoptosis.

into the cytoplasm of

Chondroitin-6-sulfate
18

Glucuronic
acid

0(1

*3)

Heparin

ing with nerve conduction due to a lack of insulation (see the figure below for a schematic representation of the myelin sheath in the central nervous system). The myelin sheath is composed primarily of phospholipids and proteins. Tiiacylglycerol and gangllosides are nor
found ln the sheath.

The answer is A: and proteins. The woman is experiencing the symptoms of multiple sciero_ sis, a demyelinaring disease. In this disorder, the myelin sheath around netl,es degenerates, eventually interfer-

phospholipids

NHS03-

Repeating disaccharide units found rn glycosaminoglycans

The oligodendroglial cells sl.rrthesize the myelin sheath found rounding the neurons in the central nervous system.

sur_

lr

tlt

i:
I

i9

The answer is B:

.'.: -:- -l: l.-l' :: --s::r..-r:.ds

: ---:, - --- :

Phospholipids.

:.

. --

--;--::

-,.: :..-.- ... 1n :' - ----lla

::----.,.: -, -:--*-JS:rg;insl its OWn ali ::i ,<,:: pr-ospnolipid complexes

The answer is B: Glycosaminoglycans. Fluoroquinoiones have been assocrated r'vith spontaneous tendon ruplure yrelding the classic histopaihologic fi.ndrngs as describe d case. Other risk factors for Achilles tendon rupture include steroid injections into the tendon, gout, rheumatoid arthritis, and renal transplantation. During

in the

-... ::- r..'urr or-ie be ing an anticardiohprn antlbody). These ...r-rb..Cre. ri'i1l bind to proteins involr'ed in coagulation
:.:-Ld Lncrease

the risk of blood clots. Antlbodies directed

:.qainst s-tochrome c, DNA, RNA splicing proteins uhrch occurs rn SLE), or ribosomes are not observed
ri1

thls disorder.

tendon degeneration, glycosamlnoglycan synthesis is increased in the extracellular matrix material of the iendon. Cholesterol, HDL, and tnglyceride have no function in the tendon rupture. In addition, sphlngostne rs
also not found in the extraceliular matrix of the tendon.

Chapter 17

Whole-body Lipid Metabolism

This chapter
and

trigly cuide) thr oughout tfu b o dy.

c1ui77es the student on the flow of lipids (primarily cholesterul and. _storage

molecular defect in this patient is present in whrch of the following proteins?

QUESTIONS
Select the single best answer.

(A) HMG-CoA reductase (B) AX,IP-activated protein kinase (C) Lecithln cholesterol acvlrransferase
(D) ABCl

(E) Cholesterol

ester transfer proteln

You have a patient whose blood work indicares high eler.,ated lir.,er enzymes. you place him on cholestyramine to lower his cholesterol. Cholestyramine acts to lower cholesterol by rnhibiting which of the following enzymes/pathways?

total cholesterol and

Current Amerlcan Heart Association Guidelines rndlcate that an adult male should have HDL levels equal to or greater than 40mg/dl. A necessary enzyme contributing ro HDL's prorecrlve effect is which o[ the following?

(A) HN,IC-CoA reducrase (B) Hepatic cholesterol synthesis (C) Reiease of bile salts from rhe gall bladder (D) Enterohepatic ctrcularion reabsorprion of biie (E) The production of chylomicrons

(A) CETP (B) LCAT

salts

(C) ACAT (D) AMP-activated protein kinase (E) Protein krnase A

Many clinicai labs report lipid values using a calculated value lor LDL. Thrs calculation estimates the cholesterol content ln which of the following particles under fastrng
conditions?

You have placed a patient on Pravachol pravasratin to reduce her cholesterol This class of drugs is effective due to a direct inhibition of which of the following? (A) Medium chain acyl-CoA dehydrogenase (MCAD) (B) HMG-CoA slnrhase

(C) HMG-CoA reducrase (D) Carnrtlne acylrransferase

(A) uot(B) I-ot-

1 (CAT-l)

(c) lDL
(D) VLDL

(E) Citrate iyase

A knockout mouse was created in which the ability to create conjugated bile sahs was grearly impaired. The
net result of this mutatron in a mouse fed a normal dlet ls which of the followinc'? (A) Steatorrhea (B) Elevated 1eve1s of chylomicrons

(E) Chylomicron
Statins are ineffective in lowering cholesterol levels in individuals wrth homozygous familial hlpercholester_ olemia due to which of the following?

(C) Deficiency ol B viramins (D) Reduced pH in the intestinai iumen (E) Reduced secretlon of pancreatic zymogens
A patient has enlarged orange tonsils, hepatosplenom_
ega1y,

(A) Hl,,tC-CoA reductase is resistant to statins (B) Statins cannot enter the liver ce11s (C) t-Ot- receptors are nonfunctronal (D) Reverse cholesterol transport rs inoperative in
patients

these

(E) LCAT is resistanr to starin acrion

loss ol sensation

ing ol the corneas.

in hands and feet, and cloud_ His HDL 1eve1s are l8mg/dl. The

You see a patient who has steatorrhea, with very 1ow lev, e1s of chylomicrons and VI_DL in the circulation. Circu_ lating triglycende leyels are extremely 1ow Examination

r48

Whole-bodY LlPid Metabolism

149

of intestinal epithelial cells shows lipidJaden cel1s A possible enzqatic defect leading to these findings is
which of the following?

(A) LPL (B) Apolipoprotein CII (C) MTTP (D) LCAT (E) CETP

(A) LDL (B) Oxldized LDL (C) Triglycerides (D) HDL (E) Oxidlzed HDL

*t

t", "

A type

diabetic who has neglected to take his lnsulin for a few days displays both hyperglycemia and hlpertriglyceridemla. The hypertriglyceridemia is due, in part, to which of the following? (A) Reduced rymthesis of VLDL (B) Reduced production of apolipoprotein CII (C) Increased fatty acid oxidation (D) Reduced secretion of LPL (E) Increased sy-ithesis of Bl00

Your 27-year-old male patient, with a BMI of 34, has a total cholesterol of '150mg/dl- and triglycerides of 610mg/dl. He exhibits planar xanthomas and has already had one angloplasty last year. This patient may be exhibiting a rare autosomal recessive disorder which generates a mutation in which of the followlng
proteins? (A) LPL

(B) Apolipoprotein Cll (C) Apolipoprotein E

(D) Apolipoprotein B100 (E) Apolipoprotein B48

lO" A 12-year-o1d female

presented with severe abdominal to have a markedly eievated plasma pain and was found

l.+' A 44-year-oldman displayed elevated cholesterol 1eve1s ' ' and was prescribed a statin to reduce such levels Statin
treatment has the potential to interfere with the s)'nthesis of which of the following?

triglyceride concentration (750mg/dl) A lipoprotein analysls reveaied elevated levels of chylomicrons and t'l-OL and reduced leve1s of HDL Which protein might
be defective in this Patient?

(A) Apo 8100 (B) APo B'18

(C) Apo CII

(A) Heme (B) Coenzyme Q (C) Ketone bodies (D) Glycogen t Et Dihydrobiopterin

(D) Pancreatic

llPase

(E) LCAT

.' a 57-year-old man has been taking low-dose aspirin to "'" .edrr." his risk of heart dlsease. He adds phytosterols to
his daily regime for which of the following? (A) To reduce circulating trlglycende levels (B) To reduce circulating cholesterol levels

""""

u.rulyri, reveals the presence of elevated triglycendes,

phoipholipids, and cholesterol esters' Levels of

car-

bohydrate and protein were normal Physical exam is unremarkable. A possible defect in the reiease of which of the following would lead to these results? tA) Cholecystokinin

(C) (D) (E)

To reduce endogenous cholesterol ryTlthesis To decrease insulin secretion To reduce fatry acld biosynthesis

(B) Insulin (C) Glucagon (D) Secretin

iiE Con..tning t''"'

'''

(E) Cortisol

the patient in the previous question, phytost..ols have the same general mechanism of action as which of the following drugs? (A) Atorvastatin

'"''""

46-year-old. man has been progressively having trouble breathing whlie walking. Walking from his car to his office has become difficult, and he has to stop to rest along the way. He visits hls physician, who ord.ers an angiogram, which shows blockage of major arteries leading to the heart. An lnitiating factor for the development of the blockage is which of the followlng?

(B) Ezetimlbe (C) Pravastatin (D) simvastatin (E) Metformin

,.iii "'"''

lau.tophages found

in arterial fatty streaks are often

ce11s.

lipid fil1ed and become foam

Such large amounts of cholesterol uptake into these cells is possible due to which of the following?

[50

Chapter 17

(A) lncreased activity of ACAf withln the foam ce11 (B) lncreased activity of LCAT within the foam cell (C) Constant SR-A1 expression on the cell surface (D) Upregulation of HMG-CoA reductase (E) Increased activity of the LDL receptor

indicated

a substantial increase in the level of lipoprotein (a). Such a resuit would suggest which of the following? (A) Substantially reduced risk for cardiovascular com-

plications

,18 A patient, 45-year-old

ma1e, BMI of 25, has had a hlstory of elevated cholesteroi (-300m9/dl), wtth normal

(B) No change in risk for cardiovascular complications (C) Increased risk for cardiovascular compilcations (D) lncreased platelet count (E) Decreased platelet count
A 16-year-old male presents to you wrth xanthomas on
the extensor tendons of the hand and Achilles tendon and arthritis of the knees. He has had one previous heart attack, despite normal cholesterol levels. Further analysis of his serum showed greatly elevated 1evels of plant sterols. The molecular defect in this patient rs most 1ike1y in which of rhe following proteins? (A) Apo B100 (B) Apo B48

triglyceride ler.e1s (-I25rng/dL), and HDL leveis ('t8mg/dl). Treatment with statins has reduced his serum cholesterol to 180mg/dl. The patient's father
had a similar history and died of a heart attack at age 48. A potential mutation in this patient would be in which of the following proteins?

(A) LCAT

(B) CETP (c) ABCI

(D) Apo B100

(E) LDL receptor

(c)

ABC1

l9

A patient sees his or her physiclan for continuing treatment of hlpercholesterolemia. Recent blood work has

(D) ABCG5

(E) MTTP

1;l
A\S\\-ERS
I
The answer is D: Enterohepatic circulation reabsorption

of bile salts.

Bec;.u.se .'i .he eierated liter enz\-rnes ln'er damage-'). a staiin lvould be relatively tsuggestile ol cortliarlrdi.ated in thrs patient, as a potential side effect of statins is liver damage Cholestyramine would be a reasonable alternative to statins Cholestyramrne is one of ihe "bile acid binders" and prevents the reabsorption ol bile salts. Slnce cholesterol is the precursor of bile salts, and. 95olo of bile salts are usua1l,r' reabsorbed back into the enterohepatic circulation, losing bile salts in rhe feces would require increased s)'nthesis of bile salts' thereby reducrng the levels of free cholesterol in the

-:''"1-:: 'r-\-j.:il.on. C,-,1-, :aa.-":aa -- '- -- : ' - - '' intr' tn. nttlt rh,'l-.-.-1 ' ' i.,1? ' -::r provide acerrl-CoA rn ihe cr-toplasm Tle :';'':-'-l-': '3e ,equrred to produce H\IG-Co-\ are sh'rur] l'-ri
'

bod;,. Statins work by inhiblting HMG-CoA reductase' Cholestlramine does not reduce hepatic cholesteroi interfere s1T-Lthesis, inhibit the release of bile salts, or of chylomicrons lts sole action ls wrth the production in the lumen of the intestine, where lt binds the brle salts so that they cannot be resorbed and sent back to the ltver. The enterohepatic circuiation is diagrammed below.

Liver

l-2NADPH (

+ 2Hr

/' '
'.r

_.+

Bile

I salts
.. t

Pancreas
I

..

:Stomach

HMG-CoA N>ZrunDP* redLrctase I

STEP INHIBITED BY STATINS

l'*coR-sH

Gall- I I- btadder-v-*-{ tt / common ."1 I oil" dud/ ,f / / EnterohePatic { circulation

carrylng ,bile salts

o
c
I I

lt

-o-

CHz

CH.-C

"t - OH
CHz
I

cH20H
Mevalonate

- | 'l',,.r, e5%w+
5%

Feces

The ansurer
stage
tlnr-,

of the intermedrate mevalonate TWo molecules acetyl-CoA condense to form acetoacetyl-CoA of which condenses with another acetyl-CoA to form p-hydroxymethylglutaryl-CoA (HMG-CoA) HMGCol ,yrrihrte catalyses this step Next, HMG-CoA reductase catalyzes the reduction of HMG-CoA to mevalonate. Statins (the class of drugs to which pravastatin belongs) directly lnhiblt HMG-CoA reductase, so mevalonate cannot be formed and cholesterol synthesis cannot continue' Statins do not inhibit the enzymes MCAD (required for fatty acid

is C: HMG'CoA reductase' The first of cholesteroi synthesis leads to the produc-

The answer is A: Steatorrhea. The pnmary reason for sl,nthesizing conjugated bile acids is to lower the pK" of the acid, so that a higher percentage ol the acid rn'il1 be iomzed in the lntestine. The greater a bile acid ls ionized, the more eff,cient the emulsifrcation is for the digestron

of the triglyceride. Without con1ugation wth glycine or taurine, th. pX^ of the bile salts is about 6 0; at a pH o[ 6.0, only 50% oi the bile salts will be ionized in the intestinal lumen, whlch would produce ineffi'clent triglyceri'de digestion, and the triglyceride content of the stool rvould infrease. By reducing the pK. to 4 0 (conlugated vr'ith glycine) or 2.0 (conjugated with taurine), greater than 6g'/o of the bile acids will be ionized, and triglyceride
digestion wl1l be maximal lf an lnability to conjugate the bile acids leads to lnefficlent triglyceride digestion, then

intestinal chylomicron formation will be reduced, not

152

Chapter 17
elevated (due to reduction of lipid uptake into the enrerocyte). Tiansport of the water soluble B r,rtamins into the

intestinal ce1ls is not dependent on lipld digestion, as is farsoluble vitamin absorption. The conjugation of bile
Cholic acid

acids wrl1 not affect the pH of the intestinal lumen, nor affect the secretlon of 4.nnogens from the pancreas to the intestine. The reactions involved in the con;ugation ol the bile acids are shown below.

will it

ATP CoASH AMP +PP;

o
il

+: H3N-CH2-COOGlycine

c**r--.-.,.so;;

il .--*'-.-*--".*-----.-

o
il

HO'

Taurocholic acid
PK^2

Glycocholic acid PKa4

",4,,,'

The answer is D: ABCI. The patient has Tangier disease, which is a delect in the AfP-bindlng cassette protern 1

(ABC1), a transporter in cell membranes whlch allows cholesterol efflux from the membrane into the HDL partic1e. Once inside the HDL particle, the cholesterol is trapped through esterification into a cholesterol ester.
The HDL pafticle can then return the cholesterol to the liver for further recycling. The de{ect in the patient is not in HMG-CoA reductase (required for the bioslnthesis of cholesterol), the AMP-activated kinase (a regulator of HMG-CoA reductase), LCAT (leclthin-cholesterol acyltransferase, the enz),rne which esterifies cholesterol in the HDL particle), or CETP (cholesterol ester transfer protein, a protein which exchanges HDL cholesterol
esters for VLDL trigiyceride).

The answer is B: LCAT. HDL is protective, in parr, due to its abllity to remove excess cholesterol from ce11 membranes and return it to the liver. In order to accomplish this, the cholesterol, after transport to the HDL particle via the particlpation of ABCI, needs to be trapped within the core of the HDL particle, and this is accomplished by esterification and converting the cholesterol to a cholesterol ester. LCAI Gecithin cholesterol acyl transferase) is the enz)rme that creates a cholesterol ester. The reaction, on page I53, is the transfer of a fatty acid from phosphatidyl choline (lecithin) to cholesterol, creating the

cholesterol ester and lysophosphatidyl choline. ACAf (acyl-CoA cholesterol acyl transferase) creates cholesterol esters in ce11s, but not in the HDL particles. CETP exchanges HDL cholesteroi esters for VLDL trigiyceride.

-. .:._..._-__-.. .'..'a :.. l,: t l; :-..- I S. : :r -: ::..'.. : : : -: -l i :a -l : :a l--: ! .ir( In-..i.(o. ;r.i thc I DL rr.cpt, -i -:( r.a:.--.r.... ...... Lpregulatrng nonlunctional LDL receptors irriL n..t ie:;
,--.::- - -.'.,
aa
- --

to a reductron ol LDL in rhe crrculation. so such rndiriduals are resistant to staiin action. FH is not due to r resistant HMG-CoA reductase, nor an inabilrtr ol strtins to reach thelr targei. FH is not related to re\-erse cholesterol transport, nor to LCAf. A diagram of receptor-mediated endocytosis, indicating the role of the
receptor, is shown belor,v

LDL particle

)---Y

^*cn l--(,
Cholesterol ester

9\

rVtV yv \
\mprex

Y t 9^"-q^
\

At lr'

tY')
Endosome Lvsosome

nt//t

,s4\

"F ,YU
:','^.1 ^

,, //U l'1 HC-O-C-R1

I

'-.,.,J1 f

HC

HC

lo lrr

-OH

LDL receptor synthesis -,-

HI

-o - e- o-cu,cu.il1cH.1.
o '" Lysolecithin

Cholesterol ester droplel Endoplasmic reticlrllrm

The answer is D: VLD[. Under fasting conditions, the total cholesterol measured will be the sum of the cholesterol in the HDL particles, the LDL particles, and VLDL. Chylomicrons should be ni1 under fasting conditions. The total cholesterol is measured, as are HDL and trig-

lycerides. Since the VLDL rs the primary triglyceride carrier under these conditions, the cholesterol content of the VLDL ls estimated to be 20ok that of the triglycende content. Thus, the formula for calculating LDL values is LDL = total cholesterol - HDL - t(Tcy5l.
The answer is C: LDL receptors ale nonfunctional. Statins are effective in lowering circulatlng cholesterol levels due to a series of events. First, the statins inhibit

is C: MTTP. The patient has abetalipoproteinemia, an absence of apo B-containing proteins in the circulation. This leads to 1ow chylomicron and VLDL levels. The problem is the synthesis of the chylomicrons and VLDL, both of whlch requlre the activity of the microsomal triglyceride transfer protein (MTTP). In the absence of MTTP activity, triglycerides cannot be transferred to the core particle as it rs
The answer being synthesized, leading to litt1e, if any, synthesis of these particles. The intestinal cel1s become laden with Iiplds obtarned from the diet and those which cannot be exported due to the inability to produce chylomicrons. Mutations in LPL or apolipoprotein CIl will not interfere wlth chylomlcron or VLDL synthesis; mutations in those proteins would lead to an inability to remove trlglyceride from those circuiating particles. Deficiencies in LCAT or ABCI, which are related to HDL metabolism, would not alfect the synthesrs of

HMG-CoA reductase, reducing intracellular slnthesis of cholesterol. The reduced cholesterol levels in the

celi then upregulate the sy,nthesis of LDL receptors,

which remove LDL from circulation, thereby reducing circulating cholesterol levels. Familial hypercholester-

L54

Chapter 17

chylomicrons or VLDL. A schematic of MTTP action

is shown below.
EB Lumen

ApoB-48 /;;\ fffi\ r _-*r_-_, I oarricre l__r* [ nr6e

(rro

'i".,',-/ [rr,
\ rG

\,n,.,/
Cytoplasm

__+

ro

corgi

:"JJ1X'JI$:;

Ribosome

^)\ A

LIPID

of insuiin, LPL levels are low, and the particles have a longer half-life in circulation due to the reduced rate of digestion, whlch contrlbutes to hypertriglyceridemia. If there were reduced syrrthesis of VI-DL, triglycerides in the circulation would be reduced, not increased. Insulin does not alter the rate of apolipoprotein CII production. The release of lnsulin decreases fatty acid oxtdation (promoting fatty acid sy,nthesis), but if increased fatty acid oxidation did occur, then tnglycerides would not accumulate in the circulation. Insulin also does not alter the sl,nthesis of apolipoprotein B100 in the liver, w-hich is required for VLDL ry-nthesis.

A model of mrcrosomal triglyceride transfer protern (MTTP) action. MTTP is requrred to transfer lipid to apo B48 as lt is slnthesized, and to transfer lipld from the cytoplasm to the lumen ol the endoplasmic reticulum as the particle (chylomicrons ln the intestine, and VLDL in the liver) is being sgrtheslzed.

lO

'9

The answer is D: Reduced secretion of LPL. Insuhn release stimulates the secretion of lipoprotein lipase (LPL) from fat and muscle cells such that the caprllaries infiltrating these tissues have the lipase bound to extracellu1ar matrix material. Then, as the triglyceride-rich particles move through the tissues, they bind to LPL vla apolipoprotein CII, and the triglyceride is digested and the fatty acids used by the tissues. In the absence

The answer is C: Apo Cll. A lack of apolipoprotein CII would mean that lipoprotein lipase could not be activated, and the triglycgdds in both chylomicrons and VLDL would be unable to be digested. This would lead to elevated leve1s of these particles, and a very high serum triglyceride leve1. Since VLDL is not being converted to IDL or LDL cholesterol levels are not elevated. Defects in either apo B 100 or apo B,18 would lead to a loss of either VLDL or chylomicrons, which is not observed. A defect in pancreatic lipase would lead to steatorrhea, as the dietary triglycerides would not be able to be digested. A defect in LCAf would affect HDL metabolism. but not triglyceride metabolism. An ovemew of the functions of the lipoproteins is presented in Table l7-1.

Table 17-1. Characteristics of the major apoproteins

Apoprotein Source
Apo A-1 Apo A-ll Apo A-lV

Primary

Tissue

Molecular Mass
(Daltons)
28,016

Lipoprotein Distribution
HDL (chylomicrons) HDL (chylomicrons) HDL (chylomicrons) Chylomicrons

Metabolic Function
Activates LCAI structural component of HDL
Unknown Unknown

lntestine, liver
Liver
I

17,414

ntestine
ntesti ne

46,465

Apo B-48 Apo B-'100

264,000
540,000

Assembly and secretion of chylomicrons from small bowel

VLDL assembly and secretion; structural protein of VLDL, lDL, and LDL; ligand for LDL receptor

Liver

VLDL, IDL, LDL

Apo

C-1

Liver Liver
Liver

6,630 8,900 8.800

Chylomicrons, VLDL, IDL, HDL Chylomicrons, VLDL, IDL, HDL Chylomicrons, VLDL, IDL, HDL Chylomicron remnants, VLDL, IDL, HDL

Unknown; may inhibit hepatic uptake of chylomicron and VLDL remnants

Cofactor activator of Iipoprotein lipase
(LPL)

Apo C-ll Apo C-lll

lnhibitor of LPL; may inhibit hepatic uptake of chylomicrons and VLDL remnants
Ligand for binding of several lipoproteins to the LDL receptor, to the LDL receptor-related protein (LR P) and possibly to a separate apo-E receptor Unknown

Apo

Liver

34,145

Apo(a)

Liver

Lipoprotein " little"

a (Lp(a))

Whole-bodY LiPid Metabolism

155

It

Secretin. Secretin is released from the j.ntestine when food enters, and it signals the pancreas
The answer is D:

n3

The answer is C: Apolipoprotein

E.

The pati'ent has dys-

betalipoprotelnemia, a mutation in apolipoprotein E'

to release a watery mixture of bicarbonate into the rntesti.ne, in order to help neutralize the acid present from

,u.h

ii-rat the patient exhibits the rare E2 form instead of the normal E3 form. Apolipoprotein E has affinlty for

the digestion thal occurred in the stomach' If the pH of thelntestinal lumen is too low, the bile salts will not

be be ionized, and emulsification of the dietary fats will mixed micelles to inefficient, as will be the formatlon of a1low intestinal absorption of fat components Digestion

of carbohydrates and protein is not dependant on bile salt lonization. A loss of cholecystokinin would result in no pancreatic zl,rrogens being secreted' and there would be no digestion of carbohydrates, proteins' or liprds within the intestine. A iack of insulin secreLion'

the LDL receptor and the LDL receptor-related protein and, as such, is lmportant for chylomicron remnant and IDL uptake from the circulation by the liver' With the homorygous E2 form, binding of the particles to their ,...pto.i is weak, and. the partrcles circulate longer than normal, contributing to the high cholesterol and trlglyceride 1evels seen in the circulation Only about l0o/o of
the indi.viduals who are homozygous for E2

will develop

ll,rtestinai 1umen. Cortlsol secretion also does not alter intestinal digestion of nutrients'

oi g1r.ugo, secretion, does not affect digestion i'n the

this condition, and in those, obesity (BMI ol 34) is a key factor which links the condition to the muiation' This (LPL) drsorder is not a problem with lipoprotein lipase from particles, so neither I.PL nor digesting triglycerides upo Ctt-it defective. As both chylomicrons and VI-DL aie produced, it is not a defect in either apo B'18 or Bl00 production or lunction

l2

The answer is B: Oxidized LDL. Oxidized LDL is taken up by macrophages, which evenlually turn into foam ..ttt i., the d,evelopment of an atherosclerotic plaque'

The higher ones LDL 1eve1s are, the more likely that oxidizi LDL will form, leading to plaque formation' The receptor which recognlzes and takes up oxidrzed LDL, SR-A1, is not downregulated, so the macrophage has an unlimited capacity to take up and store the oxldized LDL. Plaque formation does not occur due to form' elevated leve1s of nonoxidj.zed LDL, HDL of any
or triglycerides. A cartoon depiction of atherosclero[ic artery is shown below'
a

l4 fhe answer is B: Coenzyme Q.

Coenz)'rne Q is derived

from lsoprene units, which are produced in the pathway of iholeste.ol bioslTrthesls, after the HMG-CoA ..d.,.,ur. step. If HMG-CoA reductase is rnhibited (as it is by statins), then the production of the isoprenes is also reduced, and both Coenz)rne Q and dolichoi 1evheme' e1s couid become limiting. The bioslT rthesis of or dihydrobiopterin is not ketone bodies, glycogen,
dependent on isoPrene units.

normai and an

Atherosclerosis: The consequence of high cholesterol

t5

The answer is B: To reduce circulating choles' terol levels. Phytosterols interfere with cholesterol

When the level of cholesterol in the bloodstream is normal, arterial walls remain smooth and slippery.

absorptlon in the intestine (through blockage of cholesterol incorporation into the mixed mice11es, which are necessary for intestinal epithelial ceils to absorb di'etary cholesteioi), thereby leading to a reduction in circulating cholesterol levels. The phytosterols do not interfere wiih ttte blosynthesis of cholesterol, nor do they alter the secretion of insulln. Phytosterols are also not capabie of altering the rate of fatty acid bioqmthesis, nor do they affect cilculating tri.glyceride levels The effect of phytoste.ols is specific for the lnhibitlon of cholesterol absorption lrom the intestine

16
When cholesterol levels are high, excess cholesterol concentrales in the walls of arteries. thereby reducing blood flow.

The answer

is B: Ezetimibe. Ezetimibe reduces circulating choiesterol levels by blocking cholesterol absorptlon in the intestine, which ls similar to the mechanism of action of phytosterols Atorvastatin is

a statin. and its mechanism of actlon is inhibition of HMG-CoA reductase. Pravastatin is also a statin and works as does a[orvastatin. Simvastatin is yet another statin N4etformin is a lipld- and glucose-lowering drug whi.ch works via activation of the AMP-actlvated protein kinase and does not alter cholesterol absorptlon

156

Chapter 17

Table 17-2. Mechanism(s) of action and efficacy of lipid-lowering agents

Percentage Change in Serum Lipid Level

(monotherapy) Agent
Statins Bile acid resins

Mechanism of Action
Inhibit HMG-CoA reductase activity lncrease fecal excretion of bile salts

Total Cholesterol !15ok-60yo

Cholesterol

LDL

HDL

Cholesterol
Iso/o-1bvo

Triacylglycerols
!1oo/o-4oyo Variable, depending on pretreatment level of

!20v.-60%

!1soh-2orh

lloolo-zs% lsyo-aon

triacylglycerols (rnay
increase) Niacin

Activates LPL; reduces hepatic production of

VLDL; reduces catabolism of HDL

l2zo/o-20%

!loN-zsy"

Tlso/o-3syo

t2oo/o-5oTo

Fi

brates

Antagonizes

PPAR-cr,

!i2vo-15o/o

Variable,

Tso/o-isok

lzooto-soot

causing an increase in LPL activity, a decrease in apoprotein C-lll production, and an increase in apoprotein A-l production. Ezetimibe Reduces intestinal absorption of free cholesterol from the gut lumen

depending on pretreat-

ment levels of other

Iiprds

llO'L-lSW

tlsyo-zon

1o/n-3Yo

!5

if tri a cyl g Lycerols are high pretreat-

o/o-8o/o

ment

LPL, lipoprotein lipase; LDL, low-density lipoprotein; HDL, high-density lipoprotein; triacylglycerols, triglycerides; PPAr?, peroxisome proliferatoractivated receptor. Adapted from Circulation. Grundy SM, Becker D, Clark LT etal. 2002;106:3145-3451.

in the intestine. Table 17-2 summarlzes the action of

cholesterol lowering drugs.

t7'. The answer is C: Constant SR-AI expression on the cell surface. The macrophages take up oxidlzed LDL
using a scavenger receptor, SR-AI, which is not downregulated. This allows the recepror to remain on the ce1l surface and to constantly import oxldized LDL into the cell. The high leve1s of cholesterol in the foam cells is not due to a change in activity of ACAf or LCAf (which is found in HDL particles), nor is there upregulation of HMG-CoA reductase (which would produce more
endogenous cholesterol, which is unlikely since the cell is filled with cholesterol and cholesterol esters). Mac-

rophages do not use the LDL receptor for importing oxidized LDL. i.8

reteptor. The patient is heterozygous for a muiation in the LDL receptor (famillaI hypercholesterolemia). This condition leads to elevated LDL levels since ihere are insufficlent receptors available
The answer is E: LDL

to remove LDL from the circulation. If left untreated,

heart attacks are common in such patients before the age of 50. This condition is treated with statins, which reduce endogenous choiesterol sprthesis, thereby leading to an upregulation of LDL receptors, which aliows for normal LDL uptake from the circulation. Mutations ln LCAT (familial LCAf deficiency) are rare and do not often lead to premature atherosclerotic dlsease (although some exceptions are noted), but do lead to kidney and corneal damage due to large amounts of unesterified cholesterol present in those tissues. HDL level rn these individuals is usually less than 10mg/ dL, which is not observed in our patient. Mutations in CETP (cholesterol ester transfer protein) lead to elevations in HDL levels and would not be responsive [o statln action. Mutations in ABCI lead to Tangier disease, which would lead to a reduction ln HDL levels, which is not seen in this patient. A deficiency in apo 8100 would impalr VLDL syrthesls and would actua1ly reduce circulatlng LDL levels since there is less VLDL present to be converted to LDL. The diagram on page I57 depicts potentlal problems which result from defects in the LDL receptor.

- :-: l.l::,.:Gene 19p13

Cerebral atherosclerosis with stroke

ti7

Position 13

./
.............*
Mutation of gene 19p13

lncreased blood
cholesterol and atherosclerosis Coronary adery atherosclerosis with coronary occlusion and myocardial infarction

Chromosome 19

Defective lipoprotein receptor; cannot capture cholesterol for excretion into bile

General atherosclerosis with aodic aneurysm

Answer 18: Potential results of mutatrons wrthin the LDL receptor

19

tions.

The answer is C: Increased risk for cardiovascular complicaLipoprotein (a) is an LDL particle with a covalently

the blle duct, where they will be released along with the bile during fat digestion. ln the absence of acttr,rtlof either ABCG5 or ABCGB, the phytosterols are packaged into chylomicrons and are eventually delivered to the 1iver, where they are packaged into VLDL. While human cells cannot utilize phytosterols, their increased presence interferes with the rynthesis of cholesterol and

Iinked apoprotein A (linked to apoprotein Bl00) attached to the particle. The presence of this unusual lipoprorein pafticle has been positively correlated wlth the presence o[ heart disease. The role of this particle is unknown, but may be related to coagulation, since apoprotein A resembles plasminogen in structure. Lp (a) Ievels do not regulate the levels of platelets in the clrculation.

the normal cholesterol recycling within the affected patient. Patients with thls disorder develop premature

zo

The answer is D: ABCGS. The patlent has sitosterolemia, an accumulation of plant sterols (phytosterols) in ce1ls and tissues. Under normal conditions, phytosterols can diffuse into the epithelial ce1ls, but they are actively transpofied back into the intestinal lumen by an ABC-cassette (ATP-binding) containing prorein. ABCG5 (the other protein responsible for ph1t651616] efflux is ,tgCGB) Those sterols whlch make it to the liver are exported by the same proteins in the liver to

coronary artery disease. It has been hypotheslzed that the high leveis of plant sterols in the circulating lipoprotein partlcles accelerate the deposition of these sterols in the wails of the arteries, promoting atherosclerosis. This disorder is not due to mutations in either apo B100 or apo B48, as both VLDL and chylomicrons are s1'nthesized normally ln the patient. The defect is not in ABCI, as the patient does not display the symptoms of Tangier disease. The defect ls also not in MTTP, as a defect ln that protein leads to abetalipoproteinemla.

Chapter 1 8

Purine and Pyrimidine Metabolism

The cluestions in this chapter wilt test ones hnowledge concerning de noyo and saltage pa{ways related to nucleotide metabolism, "as well as the relevance of these pathways to lu.man disease , and the tredtment of human dirror*.
(A) Adenlne (B) Thymine (C) Uracil (D) Cytosine (E) Ribose-5-phosphate
Your 60-year-o1d female patient has psoriasis and has been treated with methotrexate for several years. She has no other medical problems and her preventive screen_
ings, lncludrng fecal occult blood tests and colonoscopy, have all been normal. She has developed an anemia. Which of the following would you expecr to find when working up her anemia? (A) A macrocytic anemia (B) A microcytic anemia

QUESTIONS
Select the single best answer.

':l':

Your 56-year-old male patient presents with intense redness, heat, and pain over his right great toe at the metararsophalangeal joinr. F1uid from thls jolnt shows
bifringent crystals. An X-ray of rhe foot is shown be1ow.
This disease is caused by the degradation of an excessive

amounr of which of the following?

(C) Thalassemia (D) Spherocytes (E) A 1ow viramin Br,

1eve1

researcher wants to deveiop a method of labeling purines with r5N for use in future spectroscopic stud_ ies. Purine synthesis will be done in a test tube using

the de novo pathway Which starting materials should be labeled wrth the hear,y nitrogen in order to maximize 15N incorporation into punnes?

oniy the enzymes necessary to spLthesize purines na

(A) Aspartare, glycine, and glutamate (B) Aspartate, glycine, and N5-formimino tetrahvdro_
folare

(C) Asparaglne, glycine, and glutamine (D) Asparaglne, gluramate, and glutamine

(E) Aspartate, glycine, and glutamine

A patient has been recently diagnosed with colorectal

cancer. The physicran treats the patient with a combina_ tlon of chemotherapeutic drugs, one of which is 5_fluo_ rouracil (5-FU). 5-FU is effective as an anricancer drug because ir inhibirs which one of the foilowing enzymes?

158

760

Chapter 18 ''.

(A) Adenine

(B) Guanine (C) Adenosine (D) Guanosine (E) GMP ,:i..4 Considerlng the patient in the previous question, the t"'''"' orur-lg" sand in the diapers was composed of which of
the following?

"

a penlcillin-allergic child

was given a sulfonamide for

otitis media. Human cells are resistant to sulfonamides

due to whtch of the followlng?

(A) Sulfonamides are speciflc for prokaryotic DNA

polymerases

(B) Sulfonamides are speclfic for prokaryotlc RNA

polymerases

(C) Sulfonamrdes inhibit a metabolic pathway not present in eukarYotic ce11s

(A) Xanthine

(B) Hlpoxanthine (C) Guanine

(D) Adenine

(D)Sulfonamides inhibit bacterial ribonucleotide reductase, but not eukaryotic ribonucleotide

reductase

(E) Sullonamides inhibit prokaryotic mismatch

but not eukaryotic mismatch rePair

reparr,

(E) Urate ::15- A 6-month-o1d boy was brought to the pediatncian

"'"",'

due to infections. Blood work shows fr.qr.r.t bacterial and r,rral the complete absence of B and T cel1s. Radiographic analysrs shows a greatly reduced th)'mic shadow. Tieatment of the child with a stabilized protein reverses the deficiencies. This protein has which of the following activides?

rt .

primary route of carbon entry into the tetrahydro-

folate (THF) pool rs via the serine hydroxymethyl transferase reaction. Whlch of the following is required to

convert that initial form of the THF into the form that can donate carbons to de novo purine synthesis?

(A) Converts (B) Converts (C) Converts (D) Converts (E) Converts

IMP to XMP
adenine to AMP guanine to GMP adenosine to inoslne guanosine to inoslne

(A) Glycine (B) FAD

(C) Water

(D) B., (E) Bu

li,* ""'"

the patient in the previous questions, which Con "rning metabolite will accumulate in the blood cells?

tIS. ' "'

l,tany anticancer drugs are given to patients in their nucleoside form. rather than the nucleotide form' Which enzyme below will be requrred in the conversion
of deoxyguanosine to dGTP? (A) Pynmidrne nucleoslde phosphorylase

(A) dUTP (B) dcrP (C) dATP (D) dGrP (E) drrP
in the last two ques';,,! Corl..tning the patient discussed lack of immune ce1ls ''''"""' [1o.,.r, one possible reason for the is inhibition of which of the following enz;'rnes? (A) ADA

(B) Deoxyguanosine kinase (C) Rlbonucleotide reductase

(D) Adenine phosphoribosyltransferase

(E) 5'-nucleotidase

(B) Purine nucleoside phosphorylase (C) Hlpoxanthine guanine phosphoribosyltransferase

(D) Adenlne phosphoribosyltransferase

(E) Rrbonucleotide reductase

Purine and Pyrimidlne Metabolism 161

:r

.TNSWERS

lJ:*xil lI itl' :;,,JH,,X'J":I#:

S:*,:."X

lntense inflammatory reaction to those crystals' The X-ray

demonstrated soft-tissue swelling over the first metatarsophalangeal .1oint and tlplcal gou[y erosion' Urlc acid is an insoluble breakdown product ofpurines (adenine, hlpoxanthine, or guanine). Pyrimidines (th)'T mne, ura.ii, u"a cytosine) breakdown to dlfferent water-solub1e products that do not crystallize. Ribose-5-phosphate is also degraded to very water-soluble products' The path-

and a functional folate deficiency results (see the hsure below). The folate deficrency then results i" u 62616cr--: anemia due to the lack of DNA sprthesis. Red cell precu:sors increase in mass but cannot divide due to rhe lack of precursors for DNA replication. As a result, larger rhan normal ce1ls are released lnto the circulation, although the overall red cell number decreases, resultlng i.n an anemia Both thalassemia and spherocytosis lead to microcltic anemia. Vitamin B,, levels would not be affected, and the normal occult blood tests and colonoscopy indicate that there is no bleeding leading to the anemia.

way of uric acid formation is shown beiow'

Pteridine

-,:#)'
RP

-il1\E-ln:"-"J z: T/YH\dt{*\-/-I I ,-,,*\*Ai,Z

Forate (F) oH oH+l L
t>r'rAopH |\llnoe. t

oH

ring

PABA

r___-___________

Glutamate

I
i.

GMP

liX, l.
loo]^ -"
;

Guanosine

"-t
o

'
.

-T*Y\,
H2N\N/"'-N' Guanine

It*,0*.-,v Phosphate

fu'J;"J"Jl"reductase

,,*\*l
HYPoxanthine
Aropurinot

ry^)-U\-dnr-il-* lo ' Methotrexate

Hl

fi+'

Dihydrofolate +

(FHz)

LTNADPH
I

dihydrofolate reductase

\ NHi*\ p

**lnlnn"o*,0u""

1&;':*'""^"^"

OH H. INlriH

Nunop*

"\l),
Xanthine
At toou ri

Hrr.r\*rHR4"
lH_,

|zY'\c'Hl:-R
TetrahYdrofolate
(FH+)

nol

Jxorn1n,n"

o*,ou'"

o |-"o'

I ll o^il^il)oUric

,*t^---t-**

PKa=5'4

The answer is E: Aspartate, glycine, and glutamine' As shown in the figure be1ow, the nitrogen in a purine ring

acid

------)

Urine

is directly derived from glycine, glutamine, and asparric acid. Glutamate, N5-formimino tetrahydrofolate, and asparagine do not directly donate nitrogen to the ring'

The degradative pathway for purines. Note how allopurinol' a drug ,r.d tJ ,r.u, gort, inhibrts the en4'T ne xanthine oxidase' which
reduces uric acid Production.

Aspartate

?o' /:'"'n"

ffi fn" answer is A: A macrocytic anemia' Methotrexate *"*=* THF

u.t, by inhibiting dlhydrofolate
reductase such that

FH+

cannot be formed (either from folate or dihydrofolate),

Glutamine (amide N)

162
,,'

Chapter 18
The answer is B: Thymidylate

synthase. 5-fluorouracil is a

th).mine analog (thltnine ls 5-methyl uracil), which, after activation in the cel1s to F-dUMB binds tightly to th)midylate s1T-rthase and blocks the enzqe from converlrng dUMP to dTMP (see the figure below). By blocking thymidine s),nthesis, cells can no longer s).nthesize DNA and will not replicate. 5-FU has no direct effect on dlhydrofolate reductase, amidophosphoribosyl transferase, PRPP slnthase, or UMP s),nthase. The figure also indicates the effect o[ methotrexate on dihydrofolate reductase.

The child has hereditan orotic acidurla, a mutation in the UMP sgrthase tha: leads to orotic acid accumulation in the unne (see the flgure below). Tieatment wrth uridine bypasses the block and allows UTB CTB and dTTP synthesis. UnThe answer dine treatment also has the beneficial effect of blocking

is

(: Orotate.

_O,rlp

t-cH.
.Hrtrt,'C(9oo-

t'

)l] o-\N/
I

HN\F

nrf

Aspartate
I

5-Fluorouracil

Carbamoyl lt-r t, phosphate

"--t-@;\
-O,.

5-Fluorouracil
:

-n,A
u -N/
I

i)

Y
thymidylate

synthase

yy5,1yt o-

ril o\r.r/
I

HN\CH'

H,N -ll

rp c--9H'
H

o,ctr'r-c\cooCarbamoyl aspartate

Deoxyribose-P dUMP

Methylene FH4

Deoxyribose-P

FHz
Dihydrofolate Methotrexate NADPH

dTMP

t
+

dihydrololate reductase NADP+

The answer is A: PRPP. The patient has von Gierke disease, a lack of glucose-6-phosphatase activiql When thls individual tries to produce glucose for export in

Orotic acid (orotate)

tht \su,

orotate
Shoseho\\bos\\-

g\utost-6-p\osp\are atc\\il$ales, x\rx:.h then goes through elther glycolysis (generatlng 1actate) or the HMP shunt pathway, producing excess
ribose-5-phosphate. The excess ribose-5-phosphate is converted to PRPB which then stimulates the amido-

transferase

/eaee |See
II

-\ HT
I

phosphoribosyl transferase reaction (the ratelimiting step of purine productlon) to produce 5'-phosphoribosy1 l'-amrne. This last reaction occurs because under
normal cellular conditlons, the concentrations of PRPP and glutamrne are significantly below the Km values for amidophosphoribosyl transferase. Any ce11u1ar pertur-

A*A"ooR-5-@
OMP
I

bation that increases PRPP 1eve1s, then,

will

increase

;'"1',fl::,i,,:

the rate of the reaction, producing purines thai are not required by the cel1. Thls leads to degradation of the
excess purines, producing urate and leading

F.o,

to gout'

The lactic acidosis assoctated with von Gierke disease also blocks the transport o[ urate from the blood into the urine, which contributes to the elevated uric acid Ievels seen in these patients. Von Gierke disease does not lead to elevated glutamine, ATB NADH, or dTTP
Ievels.

Block in hereditary orotic aciduria

- ----

-t

--

:-.,-.a..,.-,.-.

ll.

--

-,i :-. :_ --.:-s -:i :-:

:,rtlall-.'. lslharasLlLl ti' Ie d C'11 precursors

---.: ,-- -ir.

-;:r

.'i c.lTP rrnd cICTP The cr\-slals are made

--:-...at1. as

lhal is rhe compound that ls acc]-imulatlng'

.':--',i;'. th\-Ixine. ancl c)-toslne would not be s1-r-rthesized ls Yer)' soluble -r-. -'. Dalleni \\'llh ihis disorder. Aspartale -,::; i-. ould nol lorm crysrals if it rvere lo accumulale.
The answer is E: Inhibition

in additron to y-chain s,Ynlhesis ol hemoglobin, ls aiso an indr,rcrng inhibrtor ol ribonucleotide reductase. 1l ribonucleotide recluctase rs inhibited. the cells' abilit,v to generale deoxyribonucleoticies u'111 be impairecl, and DNA s1'nthesis rvi11 be hindered Since biood cells are regenerThe answer is E: dUMP. Hl.droxy-rrea,
zr rapid rate . tl-ie)'are one olthe ilrst cells:rffected inhibition ol DNA s,vnthesrs, and the result is a br, an .1..r.r-.. oi b1ooc1 ce11s in the patient. Llf the ans\vers

aled at

thetase

catf nval-, allou,ing UTP to be produced UTP inhihits the

Il.

of carbamoyl phosphate syn' Uridrne bypasses Lhe mutated step of the

int.-,1.i.rnr,.r-,ing step of the paLhu'a1', carbamol'1 phosohate s,vntherase Il, which halts the productlon ol orolic .rcid. therebi' lou'erlng the concenlration ol orolate in the urine. this is rhe mechanlsm rvhereb,v the cr,vstals

listecl. the srnlhesis ol onll dU\4P requires thr acliriLy' ol ribonucleotide recluctase H,vdroxyurea does noi interlere lvith the srr-Llhesis of N5-methl'LteLrahldrofoiate 5'-phosphoribt-'sr-l 1'-arnine, PRPP or adenos,vlcobalamrn.

no longer form. Undine is not inhibiting the

enz,vme

thar dlrectl,v forms orotale. nor does it inhrbit aspariate transcarbamoyiase or CPS-l \\hi1e addlng uridine does lrom b1-pass the regulate d step. it rs the synthesis ol UTP the uridine rhat ieads to the drop in orolale productron
The pathrvay of orotate s)-nthesis is shor'r''ri in the ansr'ver [o the previous c]uesllon.

t0

The answer is E: Purine nucleoside phosphorylase' The

chrlci has purine nucleoside phospholrlase defictencl: lvhich. for reasons noi )'et fullr. elucidaLecl specificaLllrecluces T-cell counts but noi B-ceils' Purtnc nucleolhal srcle phosphor,vLase is one ol the saLrage enzr-tnas base p1r-rs .o,-r...rt, guanosine or inosine ro the liee

fhe answer is D: Iack of thymidine for DNA synthesis' When UN4P synthesis is inhibrted, lhere are insulfrcient precursors ior dTNttP synlhesis (r'vhich is derived from the thymiclylate synthase reactlon) The iack .lUNtp "t. of dTTP (lvhtch is derived fron-r dTNttP) ieads to an inhibition of DNA
s1T'rthests

in red bLood

cel1 precLlrsors'

ieading to lhe megaloblastic anemia The mutaLron in hered,itary orotic aciduria does not affect lolate or B', meuboliim. Since this is a mutation rn a p,vrimidine

nbose-1-phosphate (adenosine is nol a substr'rle 1t'r thLs enzvme). HGPRT deficiencl' Ieads to Lesch-\rh:rn (there sy-rdrome, r'vhose slmptoms are qulte drll'erent ,, ,-,o ,rr.rr',-run. def:.cienc1'u'ilh an HGPRT defect-r '\PRT deficiencv 1eac1s to a buildup o[ :rn insoluble metabolite (2, S-clihldrorr'-adenine) that precrpitates in the kidne,v and ivili lea,l to renal fallure. ADA deficiency u'r1l lead to an immune cLsorder, but rn ADA deficienc,r'' both B and T ce1ls are deficient. An adenosine kinase deflciency has not been reporied in humans. An overvier'v of the purine salvage pathrvav is shor'vn belorl'-'

biosl'nthettc pathwa)', there is no ellect on adenine

Free Bases

Nucleotides

Nucleosides

Answer I0: Salvage ol bases. The punne bases fil,poxanthine and guanine react r'vtth PRPP to form the nucleotides inosine and guanosine
monophosphale, respectn'eIir HGPRT catal,vzes this reactlon. Aclenine forms AMP in a similar rype ol reaction caralyzed by APRT. Nucleotjdes
are cor-r"e.ted

to nucleosldes b1' 5'-nucleotidase'

b,v
G uan ine
I

Free bases are generaled from nucleosides

purtne nucieoside phosphorl'1ase (although note that adenosine Ls not a substrate of this enzl'me)' Deamination of the base adenine occurs r'vilh

--7:f--_+

HGPBT

Y
G ivl P

5'-Nucleotidase

ilan0stne

A\{P and ADA. Of the purines, onl,v adenosine can be phosphorl'1ated by adenostne kinase
directl,v back to a nucleotide.
Purrne nucleoside Phosphorylase

164

Chapter 18 xanthine oxidase, catalyzes two reactions. The first rs the conversion of hlpoxanthine (which is produced during the degradation of adenine) to xanthine and the second is the conversion of xanthine (which is produced during the degradation of guanine) to urlc acid. Thus, in the presence of a1lopurinol, hypoxanthine accumulates from the degradation of adenine and xanthine accumulates via the guanine degradatrve pathway Both of these compounds are more soluble than urate, thus alleviating the ma.1or problem in gout.

.1.f.., The answer is D: dGTP. With a purine nucleoside phosphorylase deficiency, guanosine will accumulate (see the

flgure in the answer to the previous question), which will inhibrt the conversion of GMP to guanosine via the actions of 5'-nucleotidase (this is also true for dGMP).
As dGMP accumulates, it wiil be phosphorylated to form

dGTP Concurrently, inosine wili accumulate, blocking the conversion of adenosine to inosine and also leading to an increase in dAfP levels. The combination of d,{TP and dGTP leads to inhibition of ribonucleotide reduciase in the thymocytes, leading to T:cel1 depletion. It has also been reported that the accumulation of deorlrguanosine triggers apoptosis in T cel1s, providing another mechanism for T:cel1 depletlon. Thrs does not affect the B cells rn thrs disorder. None of the other nucleotides listed (dCTB dTTB dlMB and dUTP) will accumulate in this dlsorder.

13r

The ansurer is B: Guanine. The patient has Lesch-Nyhan syndrome, a def,ciency in HGPRT acti\.ity. HGPRf utilizes as substrates hypoxanthine, guanine, and PRPf;

converting the free base to a nucleoside monophosphate (IMP and GMP). The enzyme does not utilize adenine, adenosine, guanoslne, or GMP as a substrate.
The reason for the aberrant behavior and developmental delay observed in this disorder has not yet been

t2 '

and xanthine. As shown belolv, the target of al1opurino1, the enzyme The answel

is E: Hypoxanthine

elucidated.

-T,Y\,
HrN\N.,''-T'
RP

AMP

t*.

The answel

is E: Urate. Patients with Lesch-Nyhan

[**"0.
+
IMP

gout as the free bases, guanine and hypoxanthine, can no longer be salvaged. As these bases accumulate, urate is produced in excess, leadsy,,ndrome develop severe

GMP

",4
Guanosine

VP' l.+

lnosine

f''
lt>

ing to gout. This is frequently seen in infants with this disorder as an orange sand-like compound. Xanthine,
hypoxanthlne, guanine, and adenlne are not accumulating in the urine, as these molecules are metabolized to produce urate.

*,*\*Arl
Guanine

-T\\"
\\

ov

phosphate
Ritrose-t

l"'

"T\\\*AI'
Hypoxanthine
Allopurinol
I

ov

Ribose-t phosphate

'I5-

The answer is D: Converts adenosine

to inosine.

The

patient has the symptoms of ADA deficiency, which

leads to severe combined immunodeflciency sl.ndrome.

runl<-/\ - \ Y nrul\.-N.'.

i /-o, xanthine l{\rHzoz /

ADA catalyzes the conversion o[ adenosine to inosine. IMP dehydrogenase converts IMP to XMP APRT
converts adenine to AMP HGPRT converts guanine to GMP, and there is no enzyrne that can convert guanosine to inosine in one step (guanase can convert guanine to xanthine in one step, but does not work on nucleoside substrates). The answer is (: dATP Due to the lack of ADA activity, adenosine accumulates and is converted to AMP by adenosine kinase (and deoxyadenosine is converted to dAMP). The d,{MP will eventually be converted to dAfB which accumulates within the cel1. There rs no accumulation of dUTB dCTB dGTB and dTTP under these conditions. Adenosine and deoryadenosine levels are also hlgh in the blood, as all tissues of the body release these compounds when they can be no longer metabolized, due to ADA defi.ciency This leads to accumulation of these toxic lntermediates in the lyrnphocytes, which are the tissues that manifest the clinical aspects of the disease.

oxidase

HN "J---An)"
Xanthine
Allonrrrinol

J o

lr
!t

o,
Xanthine oxidase

'i$'

f"r,o, -o-eKa=S4

o\^,A*/ H^H
uric

'T/Y\

acid

-------)

Urine

The degradative pathway for punnes. Note how allopurino1, a drug

used to treat gout, inhiblts the enzyme xanthine oxidase, which reduces

unc acid production.

..::

Purine ancl Pyrimrdrne Metabolism

165

::::'

Table

18-1.

Effectors of ribonucleotide reductase activity

Effector Bound to Overall Effector Bound to Substrate

Preferred Substrate
None
CDP UDP GDP

Activity Site
dATP ATP ATP ATP ATP

Specificity Site
Any nucleotide
ATP or dATP ATP or dATP dTTP dGTP

ADP

/Votes: Ribonucleotide reductase contains two allosteric sites. One regulates the overall activity of the enzyme (with dATP b ocking activity and ATP stimulating activlty). The other site regulates which substrate will be reduced (thus, with ATP bound to the substrate specificitv site, pvrimrdrne diphosphates will be reduced by the enzvme).

l7 The answer

is E: Ribonucleotide

reductase. The

cannot synthesize (which is why folic acid is

required

increase rn dATP, which occurs when ADA is defecttve, ieads to the binding of dATP to the allosteric actir-ity slte of ribonucleotlde reductase. which leads to the rnhibition of overall enz)Tne activity Thus, deoxyribonucleotides cannot be produced for the ry.nthesis of DNA, and cells are not capable of replication. Elevated

viramin i.n the human dlet). Via inhibition of THF synthesis, the target prokaryotic cells can no longer s),ntheslze dTMP and purlnes and are unable to grow and replicate. Sullonamides do not affect DNA polymerases directly, nor do they alter mismatch repair. Sulfonamides also have no ellect on ribonucleotide
reductase.

levels of dATP do not have an inhibitory effect on purine nucleoside phosphorylase, hypoxanthine gua-

nine phosphoribosyltransferase, APRT, or ADA. The regulation of ribonucleotide reductase is summarlzed

1n

lg

Table 18-1.

The answer is C: Water. Serine donates a carbon to THF to form N5, NtO-methylene THE Thls rs oxrdized to form N5, NlO-methenyl THII which is then hydro-

lyzed with water to form NlO-formyl THF (see the

i8

inhibit a metabolic pathway not present in eukaryotic cells. Sulfonamides inhibit the syrrthesis of THf; a compound that eukaryotic ce1ls
The answer is C: Sulfonamides

figure below). As such, glyclne, FAD, Br2, and Bu are not required for these conversrons to take place. The pathway for folate metabolism is shown below.

Hisiidine

Tryptophan

Sources ol one-carbon groups

Serine

Glycine

tl@ FH4 N5-Formimino

I

t HCOOH (formate)
I

N5,N1o-Methylene FH. Donation of oxidized carbon groups

I,

y,', )AD(')H \1 \-r l\AUl T,n

tl''-t

*ro.

dUMP:l

-IzNADH
<--- t

N5,N1o-Methenyl FHo

<-

Hzo

le

+ N1o-Formyl FHo

rMP*l@
FHz

@f.,rooAdenosine

l^ It5'
R

+
Purine biosynthesis S-adenosyl homocysteine S-adenosyl methionine (SAM, methyl donor)

Ns-Methyl FH4

Donation of methyl group

H::)
FHa

e,, K8J
FHn

lz-,ttomocysteine

1zr

- CH.

+.rMethionine

ATP

PPi, Pi

------+ Answer 19

66

Chapter

l8
pyrimidine nucleoside phosphorylase is not required -this pathway There is no deoxyguanosine klnase (ri.
only purine nucleosides that can be phosphorylated t' adenosine kinase are adenosine and deoxyadenosine APRT only works for the adenine base, not guanine. Ti. 5'-nucleotidase is not required as there are no dephcr:phorylation events in the pathway outlined.

f0.. The

answer is C: Ribonucleotide reductase. Deoxyguanoslne would be first acted on by purine nucleoslde phosphoryiase, whrch would produce guanine and deoxyrlbose-1-phosphate. The guanine would be converted to GMP by HGPRf, and the GMP phosphorylated to GDP The GDP would be reduced by ribonucleotide

reductase to dGDP, which is then phosphorylated again to produce dGTP Srnce guanine is a punne,

Chapter 1 I

Diabetes and Metaholic Syndrome

I i s ch ap t e r al s a pt r e s tnt s que s ti ons ittt' olt in g ov e r ctll
I

.,,,

pathways, which da not sytecifically -rildi'ess diabetes ar metabalic syndrome.

:i biochenrical

lt ole -b a dy metab olism, emph*sizing the inte gr ation

(A) Elevated levels of galactose in the lens (B) Elevated levels ol glucose in the lens (C) Elevated ler.els of sorbitol in the lens (D) Cataract lormalion (E) Increased intraocular pressure from hyperglvcemt.r
A qpe 2 diabetic has been taking metformtn to help regulate blood glucose 1evels. What ellect wll1 metformin also erert rvithin the muscle? (A) Reduce glucose uptake from the circulation

QUESTIONS
Select the single best answer.

Your diabetic patient has recentll'been place d on pramirntide (Symlin) to help control his diabetes. \Vhich ol the fo1lou'ing best describes the mechanism of action ol

this medication?

(A) It decreases glucose-6-phospate (B) It increases hexokLnase (C) It stimulates giycogen phosphorylase r D r lt inhibits glucagon secretion (E) It inhrbits insulin secrelron

(B) Enhance latty acid oxrdation (C) Redr-ice fattl'acid oxidation (D) SLimulate glucose release
r

f'

Enhance gluconeogencsis

Your 20-year-o1d male patient has had multiple epimor, and During tu'o ol these inlense hunger. He had one seizure. episodes, hrs blood glucose was 40 mg/dl. This patient was desperately tryrng to gel a discharge from the mrlitary, and you suspected he was inducing his s1-mptoms by doing whrch of the following? (A) Seif inlection of glucagon (B) Self inlectron of rnsulin (C) "Carb ioading" before exercise (D) Taking metformin belore exercise (E) Taking an amyhn blocker
socles of lightheadedness, sweating, fatigue, lre

Your patient r'vith type 2 diabetes mellitus is usually in good control u,ith an HbAIC of 7.1 and fasting blood glucose values between 90 and 100 mg/dl-. Hrs probiem is wlth his l-h postprandial glucose levels at lunch and dinner. A reca1l ol his usual diet reveals some type of meat, potato, broccoli, mi1k, and diet drink at these meals. Whlch of these loods is most likely responsible lor hi.s postprandial high blood giucose?

(A) Meat

(B) Potato (C) Broccoii (D) Milk (E) Diet drink

You see a 56-ys21-eld female patient in fo11ow-up after discharge from the hospital. She was [reated lor ketoacidosis and hyperglycemia and now is on basal and rapid actrng insulins. You wonder if she rea1ly has type 1 diabetes mellitus and was in ketoacidosis or has tlpe 2 diabetes melhtus and had a hyperosmolar state with lactic acrdosis. Which ol the following lab tests would help you determlne whether this patient has type I or type 2 dlabetes mellitus?

A parient had new glasses prescribed by hrs optomerrisr' Less than a week later, his prescription was inadequate and he could not see well with his new glasses. His

optometrist checked his vrsion twlce more ol'er the next u.eek and the patients prescription was different both times, His optometdst refers the patient to an ophthalmologist. What is the reason the patient is havrng such raprd changes in his glasses prescription?

L67

68

Chapter 19

(A) Insulin 1eve1s (B) C-peptlde leve1s (C) Fasring blood glucose 1eve1s (D) Random blood glucose 1evels (E) Hemoglobin AlC levels
-.7r,,, Your patient, mth a BMI ol36 and a warst circumference of 44 in., has a fasting blood glucose leve1 of I45 mfldL. One reason for the elevated blood glucose is which of the following?
release of glucose lrom the intestinal epi, thelial cells (B) Stimulatlon of GLUT4 rranspofters in muscle (C) Activation of pyruvate carboxylase

(A) Increased acriviry of lipoprotein lipase (B) Increased acriviry of pancreatic lipase (C) Substrate-induced activation of hormone_sensirive
lipase

(D) Increased transcription of colipase (E) Activation of microsomal tnglyceride

rransfer protein

I?'

(A) Enhanced

Your patient with metabohc slmdrome is in for a checkup. His HbAIC is 9.0 and his fasring triglycerides are 325 mg/dL. you prescribe pioglitazone (Actos) ro beter rrear his diabetes, but nothing else specific for the
high llpids. A month later, the fasting triglyceride levels have dropped to 155 mg/dl due ro a direct activation ol which of the followrng? (A) AMP-activated protein kinase

(D) Inhibition of liver GLUT4 rransporrers (E) Activation of proreln krnase B

(B) PPAR-y (C) Leptin

(D) Adlponectln

8..

Considering the patient in the prer.ious questlon, the prrmary energy source being used by the muscle is whlch of the foilowing (in the unrreared srare)?

(E) LKB1

(A) Glucose
(B) Amino aclds (C) Lactate (D) Glycerol (E) Fauy acids
,-9,

13,

Your type 2 diabetic patient has been taking metformin for the pasr 6 monrhs and has reduced fasting blood
glucose levels from 185 to 112 mg/dl Thls occurs due to which of the foilowlng effects of metformin?

(A) Activation of adenylate

cyclase

Considering the patient described 1n the last two ques_ tions, it is 1ike1y that your paiient is now leptin resistant. This has occurred due ro which ol the following?

(A) Activation of SMAD,I (B) Downregulation of rhe leptin receptor

(C) Activation of the insulin recepror (D) Activation of SOCS3

(B) Inhlbition ol the electron rransler chairL (C) Activation of LKBI (D) Stimulation of amldophosphoribosyl transferase (E) Stimulation of adenylate kinase

'i4-

The major, defining difference berween a rlpe 1 cliabetic and a type 2 diaberic is r.vhich of the following?

(E) Downregulation of anorexigenic factors

.1O. Sequelae of insulin resisrance in type 2 diabetes me1_ litus and metabolic syndrome is reduced secretion

(A) (B) (C) (D)

Weight

Ability to produce insulin

LDL levels
Blood glucose levels
leve1s

of insulin rn response to increases in blood glucose. lnsulin release from the pancreas appears to be depen, dent upon increase in concentratlon of which pair ol
metabolites?

(E) Serum triglycende

lE

(A) ATP and CO, (B) ArP and NADH

(C) ATP and NADPH

Your type I diabetic patienr was managing their disease using a combinarion ol Humulin R and Humalog. The Humalog is more rapid acting than the Humulin R due to which of the foilowing? (A) Humalog is taken orally, rarher rhan subcuraneously

(B) Humulin

R is compiexed

(D) Glucose-6-phosphate and CO,

with zinc, which slows its

absorptlon

(E) Glucose-6-phosphate and NADH

(C) Humalog is complexed with manganese, which

accelerates its absorption

tl.: '

An increase in serum free fatty acid levels, as er.rdent in individuals exhibiting metabolic sl.ndrome, occurs due to which of the following?

(D) Humulin R js raken ora11y whlch slows its absorption (E) Humalog is raken through an insulin pump
mechanism

15

: r
t' , -,-

-,--.,_.---:,:1-,.,

--- -- ,r i. ^.

l-

,,---

::,::

'

:::rsl:
-:L '-:.'..

Drsu.ro.-

:.-:::

::tt-l.l:,.n is pre\ented

it-i

Hurnalog

rD) A\lP kinast-tnduced actirillit-rfl tr- GL.'lporters

::.-':-.

and ts present in Hun.rulin R ,C) The amtno acid secLuence is completely reversed ln Humalog as compared to Humulin R

(E) Actrvatron olmuscLe acenl-CoA

carbtrrvirlse

-'

(D) The positron of two amino acids is reversed in

Humalog as compared to Humuhn
R

t9

A pregnant patient has developed gestarional dialltL.-.

One ol the consequences of gestational diabe tes t= i. -.macrosomia. Whrcl-r of the io11or'vlng is the mechanisn that causes these large for gestational age babres?

(E) Humulin

R contains genetically engineered

histidlne

residues so 1l can compiex rvrth nickel, r'vhich is not

present in Humalog

t7

Tl.p.

po11-r,rria,

diabetlcs, prior to diagnosis. display polydipsia, and poiyphagia. The po11-uria ls due lo whlch

ol the follou'ing? (A) lnsulin stimulation of urea production (B) Osmotic lmbalance due to elevated ketones in lhe
biood

(A) (B) (C) (D) (E)

z0

The anabolic elfecs ol glucose The anabolic effects ol insulin The anabolic etfects ol glucagon The anabohc e llects ol grow-ih hormone The anabolic elfects of th,vrord hormone

(C) Osmotic lmbalance due to reduced glucose levels in

lhe urine

(D) Osmotrc imbalance due lo incre:rsed glucose levels in the urlne

A patrent r'vho had gestational diabetes has .lust delivered a 10 lb baby The baby appears "iittery" and a heel stick glucose is 30 mg/dl. Which ol Lhe follourng mechanrsms is the explanation lor the newborn's blood
glucose reading?

(E) lnsulin srimulation of glucose resorption in the

kidney
18

rAr I hr molhcrs rclat.irc h1 pcrrn:ulinelnia (B) The babl's relative hyperinsulinemia (C) The mothers hlperglycemia
'
D

The polyphagia obsen.ed in the untreated type 1 diabetic, rvho has lost 6 1b in the last 2 weeks, is due to li'hich of the following?

r The babl s

hy

pergll cemia

(E) Placental insulin production

170
,1'

Chapter 19
raise glycogen slores for more glucose availabilrty durrng prolonged exercise and would not lead to hlpoglycemlc episodes. Metformin blocks liver gluconeogenesis during the fastlng state, so more fatty acids are utilized' It also reduces insulin resistance. It does not stimulate insulin release and does not produce hypoglycemra

ANSWERS
The answer is D:

It inhibits

glucagon

secetion. Pram-

lintide ls an amylin agonist used to lower postprandlal blood glucose. Amylin is a peptlde hormone secreted by the beta cel1s of the pancreas (with insulin), and inhibits
glucagon secretion when blood glucose 1evels are elevated after a meal (thus aiding insulin actlon). Glucagon stimulates release of giucose from glycogen and further raises blood glucose. Insulin stimulates glycogenesis and storage of glucose whtch lowers blood glucose. Inhibiting insulin secretion would worsen the problem ol high blood glucose levels. Decreasing glucose-6-phosphate or stimulating hexokinase or glycogen phosphorylase would increase glycogenolysis and raise blood glucose, which is opposite

fhe answer is (: Elevated Ievels of sorbitol in the

lens.

what one wants to accomplish in a diabetic patient

Sorbitol s1T-rthesis from glucose in the po1yo1 pathwa)' occurs i.n the lens of the eye. Aldose reductase converts glucose to sorbitol which then accumulates in the lens' Sorbitol dehydrogenase can convert the sorbitol to fructose. whrch can also accumulate rvlthin the 1ens. 1n d,rabetes mellitus, fluctuating levels of glucose lead to fluctuating 1evels of sorbitol, which change the consistency of the lens and therefore the glasses prescrlption Glucose and galactose by themselves do not directll'
affect the 1ens. Chronicallll high glucose and sorbitol

,'Z'

fhe answer is B: Self injection of insulin. Thrs patrent could inject exogenous insulin lo simulate an insulinoma. The s)rynptoms and lab f,ndings would be identlcal unless a C-peptide analysls was done lnjecting insulin beiween meals leads to hlpoglycemla as the insulin stimulates glucose transport from the blood into the peripheral tissues, in the absence of dietary glucose' The flgure below compares the elfects of hypoglycemia (what is occurring in this case) versus hlperglycemia (as j.n an untreated diabetic) on a patient lnjecting glucagon would, cause release of glucose from glycogenolysis (and gluconeogenesis), resulting in a higher blood glucose level. Amylin is a compound which blocks the action of glucagon, so an amylin blocker would be the same as injecting glucagon (blocking amylin activity would increase glucagon acti\1ty, since amylin is no longer active). Carbohydrate loading is an attempt to

H_C
I I

//

H-C-OH HO-C-H

H-c-oH
H-C-OH
I I

cH2oH
D-glucose
NADPH

+H*
NADP+

Aldose reductase

CH"OH

H-C_OH
HO- CI
I

t'
t

Harir'til
Sttd*y

lf} ltro{&h llffi

F.fi***{rfitiffon
fafiq$r
Bh.Eetd YlricE

H-c-oH H-c-oH
I

cH20H

&dt ty
Can!rrteit

Sorbitol (polyol)
NAD+

Egi" l;*ror i*r*ttrti! it*ud

rnlft EdlY! :ft6n {},rrd ?*lins lao rffictr di*bdtc
Bdng nrcdlciac Alcair*l

g6lL tae rlr$ah

E{*n$

l6s *6:v+ttlr1 u}rEl

NADH

Sorbitol dehydrogenase

Inl&r$ t* t*d6 dltirtG* Ersdklffi

+H+
CH"OH

C=O
t$-16g t*rhorrYdflB
t

t'
I
I

. 2*t$A*tP**{l . O.i r.]p{* 0t} frsitit.dre

fdgr.#!* SrG.c$E{d ttqhni*} ' &atr*t*rerclta - or* lsso*lyccBdt

" IIi$rll*t

HO-C-H H-C-OH

' *.S.ry t{ cl

ro{*{not dk*} . $-5pl6ct*lflrdatErdy

rS6c&l eat$*ha{t$elt

H-c-oH
I

cH2oH
D-fructose

171
::l:l: .-:l:;-'-.-:t '':::' '11' '--:l--:---::" --- :-111: ' --:'

lltr--

illii-i,r>i -l- -:-'.- - ----': ::'ssl1r' ho\\'e!e r' the con\-erpathrvai' ol >ion ol glu.cse to sLrrblt;1 u.r1l The po1vo1
sugar metabohsm is shor'i''r-r on page 170'

cirares. Broccoli h.ls i11ore ioElpL-\ ''ii-i-r''l' "r'r--iS ':l- 'a much lolver gl1'cemrc inder' \leat is moslh'proii-r': 1ou-er gh-and fats. Mtlk contarns proleins and so has a

The answel is B: Enhance fatty acid oxidation' proformin, through its activation of the AN{P-activated stimulate glucose enir,v lnto the muscle tein kinase, will (thus. ansu'er choice A is rncorrect) and also increase kinase iatry acid oxidarLon. The AMP-activated protein and inhlbit acetvi-CoA carboxl4ase .r.L1i phosphorylate decarand rvr1l piosphorylate and aclivate malonyl-CoA ma1on1'1-CoA levels drop' leading to boxylase. Thus, and enhanced entry ol fatty acicls into the mitochondria' latty acid oxidation This occurs as the an increase ln malonyl-CoA inhrbitron ol carnitine palmityl transferase I is nor,v hfted due io the reductton ol malonyl-CoA (see the figure belolv) Metformin does not slimulr''let1eve1s

has lats and cemic index lhan potaloes (w-ho1e mrlk also Diet drinks conrain no an even lower glycemrc index) index carbohydrates Jr caiories. A table of the glycemic in Tabie 19- i , r'vilh the values of .o*-on foods ls lisred adlusted to a lvhile bread leve1 of 100'

Table 19-1.
Breads Whole wheat
Pumpernickel (whole grain
rye)

Cookies

100

Oatmeal
Plaln water
100

88

crackers

Cereal Grains
BarleV (pearled)

Fruit
Apple Apple juice Orange
Raisins

muscle lacks late glucose ,elea,e lrom the muscle ' as the also reduces glucose-6-phosphatase activity Metformin in the liver at a rranscriptional level'

!1r.or-t.og.r-t.sis

36 Rice (instant, boiled 1 min) 65 81 Rice, polished, (boiled

10-25 min) Sweet corn

cPT-1
Mitochondrial matrix

Fattv-acvl CoA

a\
./

AMP+

Root Vegetables
Potatoes (instant) Potato (new, white,
boiled) 120

Dairy Products
lce cTeam

Malonyl

CoA'

TF*Lql:
Acc.2

fi)\

AcetYl CoA'

{ c

PK

80
/

Whole milk

Potato chiPs
Yam

Srlm

mllK 52

Yogurt

Regulation

of fattlr acl'l-CoA entry inlo muscle mllochondria. to iri e..tyl coA carbory'lase-) (ACC-Z) converts acetl'l-CoA palmitoyi translerase 1 (CPT:D' -4""1'it"a, u'hich rnhibrts carnltinetnto the mitochondria (2) Howentry
,f-r.r.Uy

Legumes
Baked beans (canned)
10

Sugars
Fructose Glucose Lactose Sucrose
138

.r.r, n, .rr"rgy i.r'.l,

of drop, AMP ievels rise because of the activity (3) The increase in AN'IP levels actrvales rhe adenyhrJk nase reaction (AMPK), w-hich phosphor,vlates and the AMP'actir.ated protern kinase and also phosphorylates and activates malonyl-CoA inactivares ACC-2, a.**"t-f'C"4, (MCoADC) The Jecarboq'1ase converls malonvl-CoA ^* ih.r.by relieltng the inhibrtion of CPT-I and allor"'ing ^."yf a"o enlry lnlo the mitochondria This allorvs the muscle to fr,;:;;t', generate ATP r.'ia the oxidation ol fatty acids'

frf"."'*g

farty acyl-CoA

Butter beans
Garden peas (frozen) Kidney beans (dried) Kidnev beans (canned)
Pea n

46 85 43
15

57 83

uts

Pasta Spaghetti, white, boiled

Breakfast Cereals

61

All bran
Cornf lakes
121

Ihe ansurer is B: Potato' Amounts of simple carboindicators of hydrates in a meal are the most reliable As the HbAIC nears postprandial nse in blood glucose

Muesli

values are normal or target values, fasting biood glucose glucose values have a usuaily nor*.-^l and postprandial Proteins' much more rmportant effect on the HbAlC are ahsorbed more fats, and .orrp1.* carbohydrates can be vier'ved s1owly than simple carbohydrates This (the abrlity of a food to rapidly as the glycemic index the glycemic .aise biood glucose) A higher number for hlgher rise of blood g1uindex means a more rapid and index of the cose. Potatoes have the highest glycemic

6 Ihe answel is B: ('peptide levels' lt is difficult' at and times. to differentlait typt I diabetes mellitus
lar stale and lactic acidosis unless testlng for
nr-rd,ro,

hyperosmoketoacrdosis from type 2 diabetes mellitus B-hydrory.buqrate ordered ujhen acidosis is noticed At rhls polnt
(ketone bodies) are specifi
ca111

acetate

in

the

L72

Chapter 19 patients disease process, ketone bodies should be normal. The patient is already on insulin, so rnsulln 1evels the f,gure below for an or.erview of insulin biospI drabetes mellitus (no endogenous insuh: produced) should glve a very low or nonexistent ler-e of C-peptide, whereas tlpe 2 diabetes mellitus (insu, 1in resistance) should give a normal or hrgh 1eve1 o, C-peptide. Exogenous (commercial) insuhn lacks th.C-peptide, so the lnjecred insulin w111 nor interfere w.ith this measurement.
thesrs). Type
(see

would not be helpful. Measuremenr of biood glucose levels, whether fasting or not, and determ.ination of HbIAC levels cannot differentiate rlpe I from type 2
diabetes (since the patient rs on insulin). Since insulin is secreted as a macromolecule and does not become active

until C-peptide is cleaved from the macromolecuie, a C-peptlde 1eve1 would be helpful ln this differenriarion

tr

chain

chain

?F S

B ==_.'l'=
secujnce

chain

A chain

jr

1, ll
lJ

,i
I

Endoplasmic reiiculum

....-V.NHg*

----+

Golgi apparatus

Preproinsulin

vl

li ff
u
lnsulin

n
.-...-

Signal sequence

/ )

Proinsulin

C-peptide

Panel A indicates the amino acid sequence of mature insulin, whlle panel B indicaLes rhe sreps involr,ed in converting preprornsulin to mature rnsulin. Note the release of the C-peptide as insulin is converted to its mature form.

7.

of pyruvate carboxylase. The patient most likelyhas metabolic syndrome, and rhe liver has become resistant to the action of insulin Because o[ this, gluconeogenesis is enhanced in the liver. Due to the large size of the adipocytes, free fatty acid ievels rn the portal circulation are high (visceral adipocytes release compounds into the portal veln), and the liver is oxidlzing fatty acids for energy, generating acetyl-CoA. The acetyl-CoA ls an activator for pyruvate carboxrylase, and an inhibitor of pyruvate dehydrogenase. The activarion of pyruvate carboxylase contributes to the activation of gluconeogenesis, and high blood glucose levels. The intestinal epithelial ce11s release glucose, obrained from the diet, down its concentration gradient, so enhanced release would only occur when blood glucose 1eve1s were very low. If GLUT4 transporters were stimulated in muscie, blood glucose levels would drop, as the muscle would be capable of removing glucose from the circulation. The insulin-resistance that is occurring also affects
The answer is C: Activation

glucose transport into that tissue. Liver utilizes both GLUT1 and GLUT4 transporrers, but due ro the insu1in resistance, the liver is exporting glucose rather than using it. Actll,ation of protein kinase B occurs when insuiin blnds to its receptor; since the liver is resistanr to insulin action, the level of protein kinase B aciivarion
is reduced.

iting

The answer is E: Fatty acids. Since the parienr is exhibsrgns of metabollc sl,ndrome, a key component of

which ls insulin resisrance, rhe muscle has difficulty in transportlng glucose from the circulation lnto the tissue. Thrs means that the muscle will use fatty acids as its primary energy source. When abundant fatty acids are available, the muscle will utiiize rhe fatty acids preferentially and will not use amino acids as an energy source.
The muscle (other than the heart) does not utilize lactate for energy and glycerol can oniy be metabolized in the liver, as that is the only tissue which contains glycerol kinase. The use of fatty acids by the muscles, instead of

the muscle, such that insulin can no ionger stimulate

l,
1:-: :

The answer is C: Substrate.induced a<tivation .l,r'.-a -l

of hormone.
---

The answer is D: Activation of

S0(53. ,. -. -,i :..:-'.e s

Signal

n-{:l .r'11-il -1-'.--.'ll a- '-:,r . aJ-'-*a l- : lj

aItas. HLrrrrLrne sonsitile itpase has a irr\\

'a;1:,r,

-.,--.

i:la jl'.1.-,.,'.,.::-,-. -,: -. l.--:: :.- :.C.plOrS $hiCh

itf in the absence ol actir ation b\ phosphon-l:rrrLrll . i: protein kinase A. Il 1.ou combine the hrgh sr-rbsrrare
(triglycende) r,vith Lhe 1or,v 1er.e1 ol activit)' ol hormone-sensrtive hpase , one sees a release ol fatt1, acrds from the adipocyte u.hrch is greater than normal (r.r.hen triglyceride leve1s are lolv). The increase in free latty acid ler.els then aids ln promoting insulin resisrence rn both the adipoci.tes and muscle ce11s. This occurs because the muscle and fat ceLls u'i11 use fart). acids as an energy source instead ol glucose, therebl, contributing Lo hrgh blood glucose ler-els. The increased serum free latty acid 1el'eis do not alfect the acrivit,v ol lipoproiein lipase (ivhich removes fart,v acids from VLDL and chylomicrons), nor pancreatic lipase (which cligests dretary trigll,cerides in the intestinal lumen). The lart,v acids do not aflect the transcription rate of colipase. and while the 1er-e1s ol fattl' acids u,jthin the iiver ma1, alter the activLLl' of microsomal triglvceride transler protein (MTTP). an alteration in MTTP actir.it)- does not lead tct Lhe increase in free fattr,acid levels in the blood.
1eve1

the release .-r .,:r:,:-:lsenr. n.uropeptides, i,r,hici-r signal ''stop earrng Hou'ever. leptin aiso induces ihe expression ol suppressor ol c).tokine srgnahng (SOCS3), which blocks the action of leptln. With constant leptin release, SOCS3 levels are raised, and Jeptin can no longer induce the release ofanorexrgenic signals, leading to overeating. The Leptin pathwa), (a JAK/STAT receptor and signaling mechanism) does noL invoh.e SN,IAD'+ (a component ol

the TGF-B signahng pathrval). The leptin resistance rs not due to dolr,nregulation ol the leptin receptors, or the activation of the insulin receptor. The dou,r-rregulation
of the reiease of anorexigenic lactors ls a consequence ol

leplin resistance, noi

a callse.

10

The answer is C: ATP and llADPH. The B-cells

ol the

pancreas monitor both ATP and NADPH levels in order lor insulin release to occur. The NADPH leveis are increased through enhanced p),ru\.ate cycling (see the figure belor,ii, r,vhich occurs r,vhen pyruvaie le\-e1s increase (w'hrch is correlated r,vith an increase ln glucose levels r,vithin the F-ce11). Increased giucose also leads to an increase in ATP, rvhich leads to changes rn ion fluxes across the membrane, resulting in insulin release. Glucose-6-phosphate, carbon dloxide, and NADH are not necessary for insulin reiease in response to glucose.

12

The answer is B: PPAR-y. Thiazolidinedrones (TZDs),

ol which proglitazone is a ntember bind to peroxisome prolil'erator actlvated receptor-y (PPAR-y) in the adipocl.te and acLir.ate the slnthesis and release ol adiponectin, u,hich acts on target ce11s tit reduce blood glucose Leveis (b,v upregulating GLUT4 content of tl're membranes) and [o reduce circulating triglyceride levels (through phosphorl lation and inhibition of acetyl-CoA carbox;,lase 2, r'r,hich reiieves the inhtbition of carnitine palmitoyl transferase 1) While adiponectin 1evels rise, rvhich leads io a stimulation of the AMP-acri\,ated
Mitochondrion Cytosol
Pyruvate Pyruvate

Cytosol
Glucose

Answer 10: Pl.ntr.ate cycling: An increase in

pancrealic glucose levels leads to an increase in p),mvate, rvhich enters the mitochondna.

I Pyruvate
Prruvate

tvtatict--+ trtROpU
enzyme I

The pyr-uvate can be converted to oxaloacetate, then malate, which re-enters the cytoplasm and rs converted back ro pyrur.ate ua malic enzpre, generatlng NADPH in rhe process. Conr.ersel)', p)'1-uvate can be conr.erted

*",,.f,*roorn
"nrrr"
I

/ \ eoH carb6xylase/ I Acetyl CoA /

Malate

/-'--\
OAA Malate

I I OAA Il
I

Fatty acids

4
CoA

lo

Malate

<-

//

\r+Acetyt

acetyl-CoA, generate citrate, whrch can then leave the n'ritochondna and regenerate p1t-uvate, also generating NADPH rn the reaction qcquenr e. The pln, reas llsn cnntain. a c) l.r

Citrate
\
V
I

____>

Citrate

Fumarate

socitrate

lsocitrate

plasmrc, NADPT dependent, isocitrate deh),drogenase, r'vhich provides a third mechanism for raising NADPH 1eve1s rvhen pyrur.ate 1erels are increased r'r,rthin the pancreatic B-ce1l.

Succinate

/ t"v o"n"oJ33:i,l'#: F"* NADPH

cr-KG

----->

+-

C[-KG

Succinyl CoA

^/

L74

Chapter 19 concentration of the rnsulin has to be reduced (through dilfusron) for monomers and dimers of insulin to leave the zrnc complex. This dramatically slows the time ol rnsulin appearing rn the circulation. Humalog, with a slightly different amino sequencc, is not complexed with zinc and ls absorbed much more rapidly lrom the injection site than Humulin R. Both types of insuhn are taken subcutaneously, not oraily, even u'hen using an insulln pump. Pumps will not alter absorption, just delivery and time of dehvery of the insuiin. Humalog is not complexed wlth manganese.

protein kinase , neither of those effects is due to a direct interaction with the TZD. LKB1, an upstream kinase responsible for activating the AMP-actlvated proteln kinase, and leptin are not involved in the response to TZDs. The structure of Actos (pioghtazone) is shown
below.

oY*Yo
HzC

FS
P
CHz

The thiazolidinedione pad of the molecule

a
HrC

t6 The answer is D: The position of two amino acids is reversed in Humalog as compared to Humulin R. The
sequence of Humulin R is the same as natiYe insulin, but Humalog has switched the posrtions of amino acids 28 and 29 of the B chain (see the figure in the answer to question 6 of this chapter) and is designated as lyspro insulin, for the two amino acids that have sr.vitched positions (regular insulln has the sequence pro-1ys at these positions, whereas lys-pro insuirn has iys-pro at these positions). The C-peptide is removed from both

()
HzC

*J

CHs

Pioglitazone

forms of insulin (insuhn would have no biologlcal activity if the C-peplide were not removed), and both
contain the same disulfide bonds. There are no his-tags presena on Humulin. This minor difference in rnsulin structure allows Humulin to be absorbed much faster than Humalog, yet stil1 retain its normal affinity for the

|3. tne answer is B: Inhibition of the electron transfer ' chain. Metformin partially inhibits complex 1 of the
electron transport charn. This leads to reduced ATP production, which, as energy is required, rapidly rncreases AMP leve1s due to the adenyiate kinase reaction. The increase ln AMP levels leads to the activation of the AMP-activated protein krnase (AMPK), which is the pri-

insulin receptor.
17

not actlvate adenylate cyclase, nor does LKB1 (it has been postulated that LBKI is constantly phosphorylating the AMP-activated protein kinase, but a phosphatase is always inactivating the AMPK. When AMP levels nse, however, AMP inhibits the phosphatase, ieading to fuily active AMPK). Metformin also has no direct effect on the rate-limiting step of purine production, amldophosphoribosyltransferase, or o[ adenylate kinase.

mary messenger for metformlns effects. Metlormin does

The answer is D: 0smotic imbalance due to inqeased glucose levels in the urine. In an untreated type 1 diabetic, glucose ler,els in the blood exceed the renal threshold for reabsorption of the glucose from the urine, so blood glucose levels rise in the urine. This creates an osmotic imbalance, whrch forces more water into the urine, leading to po11'uria (frequent urination). This is not due to urea production (and, since a type 1 diabetic does not produce insulin, insulin cannot be stimulatrng urea production). It is not due to an increase of ketones in the blood. And, since insulin is not present, it cannot be due to insulin stimuiation of glucose resorption in
the kidney.

|4,,

Ttre ansvuel is B: Ability to produce insulin. By defrnition, a t)?e 1 diabetlc cannot produce insulin. A type 2 dlabetlc produces insulin, but has become resistant to the effects of insulin. Werght does not differentiate bet-ween

tlpe 1 and 2 diabetics (aithough most type 2 diabetics

j.n both are overwelght). Btood glucose levels are elevated

types of di.abetes and cannot differentiate between them. Neither LDL levels nor triglyceride levels wrli differentia[e between these two ma]or forms of diabetes.

-15,. The answer is B: Humulin

R is complexed

with zinc, which

slows is a hexamer complexed with zinc. After injection,

its absorption. Humulin R

(regular acting)
the

*8,. The answer is B: (ortisol stimulation ol amino acid release from the muscle. The weight loss seen in tlpe I diabetlcs (untreated) results from the need of the Iiver for gluconeogenic precursors, many of whlch are derived from amino acids obtained from muscle prolein breakdown. Cortisol release w111 signal the muscle to release amino acids for use by the 1iver. Glucagon signals triglyceride degradation, not productlon. Insulin does signai an inhibition of fatty acid oxldation, but that is not occurring in an indivrdual who does not make insulin (type t diabetic). Since the muscle is

Dlabetes and Metabolic Syndron-re 17 5

oxidizing fatty acids for energy, there is no activation of the AMP-activated protein ki.nase, as the energy levels are not 1ow Muscle acetyl-CoA carboxylase 2 (which produces malonyl-CoA, which rn'ou1d inhibit fatty acid oxidation via inhibiting carnitlne palmltoyl transferase 1) is not activated under these condrtions, due to the lack ol insulin.
19

of hyperglycemia), and a fivefold increase rn strllborn rates are all complications of uncontrolled gestational
diabetes. The answer is B: Ihe baby's relative hyperinsulinemia. During pregnancy, the fetus is oversupphed with glucose from the mother causing the fetal pancreas to

The answer is B: The anabolic effects of insulin. In pregnancy, the placenta preferentially shunts glucose to the developing fetus. This, along with placental hormones, causes a functional "insulin resistance" in the mother. Because of the higher glucose level in the fetus, the fetal pancreas produces more insulin. lnsuiin is the major anabolic hormone of the body stimulating glucose uptake into the ce11s and stimulatrng extra growth. Glucagon, growth hormone, epinephrine, corticosteroids, and thyroid hormone are all catabolic hormones that counter insulin and stimulate glucose release from the ce1ls to increase blood glucose. Glucose ltself is neither anabolic nor catabolic. Large for gestational age babies, dehydrated babies (from the osmotic diuresis

overproduce lnsulin. At delivery the glucose supply from the mother is suddenly terminated and the relative hlperinsulinemla of the baby causes hlpoglycemia until the babys body can adjust to this new enlT ronment by decreasing insulin release and increasing glucose release. Hlpoglycemia in the flrst few hours of the newborn's life is a common complication of gestational diabetes. Newborn hyperglycemia wouid not give a heel stick of 30 mg/dl of glucose. The placenta does not make insuhn and the insulin molecule cannot cross the piacenta, so the mother's relative hyperinsulinemia is not the cause of this problem. While the mothers hlperglycemia has 1ed to the baby's reiative hyperinsuhnemia, the mother's blood glucose 1eve1s do not cause the drop

in the baby's blood glucose levels after birth.

Chapter 20

I{utrition
and Vitamins
This chapter covers cofactors denved from vitamins, as well as the basic concepts of biochemical nutntion.

QUESTIONS
Select the single best answer.

A patient has been diagnosed with abetalipoproteinemia. A possible deficiency in which of the foilowing vitamins could occur ln this patient? (A) Vltamin B, (B) Vitamin B,
(C) Vitamin C

just initiated "1r.. Consider a 4O-year-old man who has

(l-l\ Vrtamin

24hfast. Which of the followlng cofactors are necessary so that his blood glucose leve1s can be kept constant? (A) 86, biotin, and niacin (B) Bo, niacin, and vitamin D (C) Bo, blotin, and vitamin D

(E) Niacin
A deficiency in which of the followlng vitamins will lead to a functional folate defi.ciency?

(D) Biotin, niacin, and B,

(E) Biotin, niacin, and vltamin K

::2.

laboratory study, volunteers were made experimenta11y vitamin Bo defici.ent, and much to the investigator's surprise, a mild hypoglycemia, wlth ketosis, was noted. The hlpoglycemia is a result of which of the following? In
a

(A) (B) (C) (D) (E)

Thiamlne Niacin Riboflavin

B,,

Vitamin

(A) Inhibition of phosphoenolpyruvate carboxykinase (B) inhibition of pyruvate carboxylase (C) Reduced actlvity of glucose-6-phosphatase (D) Reduced activity of liver glycogen phosphorylase (E) Reduced insulin secretion

A woman, who eats a standard meat-containing diet, has had one child born with a neurai tube defect, and is considering becoming pregnant again. Blood work showed norfLal levels of B,, and total folic acid (specific type of folic acid not specified). One possrble explanation for the woman's difficulties in her first
pregnancy is a thermolablle variant of which of the fo1-

lowlng enzymes?

,. i

An individual who has been on a ionS-term diet will

have reduced the transcription of which of the following
enZ\'rneS?

(A) Malic errzyme

(B) (C) (D) (E)

Carnitine acyltransferase 1 Carnitine acyltransferase 2 Medium chain acyl-CoA dehydrogenase Fatty acyl-CoA s)'nthetase

(A) N5, N1O-methylenetetrahydrofolate reductase (B) Serine hydroryrmethyl transferase (C) Ornithine transcarbamoyiase (D) Phenylalanlne hydroxylase (E) Tyrosine aminotransferase
Concerning the woman in the previous question, alteration of her diet in which of the following ways would be beneficlal for her future pregnancies? (A) B,, supplementation r B) Folate supplementation (C) lron supplementatlon (D) A meat-free diet (E) Homocysteine supplementation

.,,. Which one of the fatty aclds listed below can be slmthe-

sized by humans?

(A) Cis L9,l)CrB.2 (B) Cis Aq, 12. I 5 Cl8:l (C) Cis 45, B, l3 C)0:) iD) Cis 45. 8. I l. 14 C20.4 (E) Cis Alo C16:l

176

Nutritron and Vitamins 177

A high-protein low-carbohydrate diet has, as its biochemlcal basis, the potential to lead to weight loss due to which of the foilowing? (A) Allosteric inhrbrtion of fatty acid sprthesis

t+

Considering the patient in the previous question, a food

that should be incorporated into the patient's diet

which of the following?

is

(B) Increased glucagon release (C) Minimal insulin release (D) Reduced cortisol release
r

lncreased urea prod uct ion

15

(A) (B) (C) (D) (E)

Butter
Margarine

Olive

oi1

Peanut oil

Coconut oil

10

The neurotransmitters GABA, dopamine, and histamrne are all derived from an amino acid precursor. The gen-

eration of these neurotransmitters from the appropriate amlno acrds requires which one of the following cofactors?

A 9-month-old child of strict vegan parents is brought to the pediatrician due to perceived muscle weakness.
Due to their strlct dletary beliefs, the child has not been given r,rtamin supplements. An image of the anterior of the knee reveals cupped and wrdened metaphyses. As the chlld is very fair skinned, the parents always cover

(A) NAD. (B) B, (C) B, (D) BO Irr B,, '

up the child when they go outside such that minimal skin ls exposed to the sun. In order to correct these
probiems the physician prescribes treatment with which

l,l A patient presents with

episodes of flushing, diarrhea, abdominal cramping, and wheeztng. His blood pressure and pulse rate are normal during these episodes. Physrcal exam is normal except for scattered telangiectasias. In order to diagnose this problem,
a

of the following? (A) Vitamrn D

24-h urine collec-

(B) Vitamin K (C) Folic acid (D) \,'itamin B,, (E) Vitamrn E r5
A patrent has had a series of blood c1ots, and has been placed on warfarin to reduce such incidents. Warfarin exeils its effect by blocklng which of the foliowing?

tion showed elevated levels of 5-H1,A,A. (5-hydroxlnndole

acetic acid). The chemical responsible for this above s1mdrome rs a derivative of which amino acid?

(A) Alanine

(B) Serine (C) Tyrosine (D) Tryptophan (E) Phenylalanine t2

An indlvidual has developed pancreatiti.s, and with it,
steatorrhea. The patient also reports problems wrth his

(A) Platelet biogenesis

tBt

(C) Clottrng factor sgrthesis ' (D) Vltamin E activity

Pho>pholipid synthesis

(E) Formation of y-carboxyglutamate

night vislon, although visual acuity appears normal.

Another expected finding ln this patrent would be which of the following?

l7

Consrdering the patient in the previous question, which food should be avolded rn large quantities while the patient is on warfarin?

(A) Nystagmus (B) Easy bruislng (C) Dermatltis (D) Loss of teeth (E) Orange tonsils r3 You are treating a patlent with
malabsorption problem, and you suggest that the patient switch his or her diet ro one that con[ains which of the followng? (A) Triglycerides with long-chaln fatty acids (B) Triglycerides with medlum-chain fatty acids (C) Triglycerides with short-chain fatty acids (D) Triglycerldes with a mixture of long-chain and short-chain fatty acids (E) Triglycendes with a mixture of long-chain and medium-chain fatty acids
a fat

(A) (B) (C) (D)

Trout

Mllk products
Green leafy vegetables

Orange-yellow vegetables
Steak

(E)

Chronic alcoholics often develop fatty and leaky livers,

as the primary site of alcohol detoxification ls the liver.

Protein and VLDL secretion can be impaired ln such parienrs due to which of the following? (A) Ethanol lnhibition of mlcrotubule formation (B) Ethanol inhibition of gene transcription (C) Acetate reduction of intracytoplasmic pH (D) Formation o[ acetaldehyde adducts with cytoplasmic proteins

(E) Increased ketone body levels within the cytoplasm

178

Chapter 20
2A

.19,. A 6-year-o1d girl has a history of seizures, which are only marginally reiieved by standard medlcations.
Switching her diet to which of the following may help her condition? (A) 80o/o far.,20ok combined carbohydrate and protein, by weight (B) 50olo fat, 50ok comblned carbohydrate and protein, by weight (C) 20'k fat, B0o/o combined carbohydrate and protein, by weight (D) 75'k protein, 25o/o combined carbohydrate and fat, by weight (E) 50olo protein, 50o/o combrned carbohydrate and fat, by weight

42-year-old woman presents with tiredness and lethargy She has tingling in her hands and feet. Blood

work shows a macrocytic anemia, along with

vated levels of which metabolite?

elevated homocysteine 1eve1s. One would also expect to see ele-

(A) Acetic acid

(B) Ketone bodies (C) Methylmaionic acid (D) Propionic aci.d (E) Succinate

Nutrition and Vitamins 179

ANSWERS
'i"'
The answer is A: Br, biotin, and niacin. Blotin ls required for pvruvate carboxyiase (the erq-me that converts p1'mvate to oxaloacetate), an enz)'rne necessary for gluconeo-

answer (carnitine acyltransferase l and Il, medrum-chain -Lthetase) acyl-CoA dehydrogenase, and fatty acyl-CoA syT are a1l involved in fatty acid oxrdation, a process which would be increased during fasting, and r'vhose enzyme 1eve1s w-ould not be decreased under these conditions.

'

genesls from any TCA cycle precursor (and py:uvate). The Bo is required for glycogen phosphorylase (the ablllty to produce glucose from giycogen) and for lransamination reactions, which are necessary for ami.no acids (such as alanlne, aspartic acid, and glutamic acid) to provide carbons for gluconeogenesis. The niacin is requi-red to produce NADH, which is needed to reverse the glyceraldehyde-3-phosphate dehydrogenase step dunng gluconeogenesis. lt is also needed for lactate lo be converted to p).ruvate, and lactate (provided by the red blood ce1l) is a major gluconeogenic precursor. The lipid-soluble vitamins (D and K) are not required for glucose productlon and release (l'itamrn D is needed for calclum metabolism,

.4.

ttre ansner is C: cis 45,8, 13 (20:3. Humans can slrLthesize fatty acids of the ro-7 series or higher, but
not of ro-6 or lesser. This is due to the limitation of the desaturase system (see the figure below-), which can only introduce double bonds at positlons 4, 5, 6 and 9,
a subslrate that contains at least 16 carbons. Answer A is an r.rl-6 fatty acrd, as are answers D and E. Answer

in

and vitamin K is required for the carboryiation of g1utamic acid side chains of proteins involved in blood clotting). Vitamin B,, is nol required for glucose production directly, and its absence does not lead to hlpoglycemra. .r2'.,, The ansurer

B is an t.:i--3 latty acid. Answer C is an rtl-7 fatty acid, and would be s1T-rthesized as fo11ows. Start wLth Cl6:0 (palmitic acld), and add a double bond at position 9, creating a cis A9 C16:1. Elongate that fatty acid by two carbons, creating a cis All Cl8:1. Desaturate this lS-carbon fatty acld at position 6, creating a cis A6All C18:2, r'vhich is elongated by two carbons, producing a crs A8Al3 C20.2. Desaturate rhis latty acld at position 5 and the fina1 product is obtained.
.r$,-.. Jhs answer

is D: Reduced activity of liver glycogen

phosphorylase. Glycogen phosphorylase requires pyridoxal phosphate (derived from litamin Bu) as an
essential cofactor. The role of Bo

'

is D: Vitamin E. Abetalipoproleinemia is a disorder in which neither nascent chylomrcrons nor nascent VLDL can be produced due to a defect in the
microsomal trigiyceride transfer proteln. Fat-so1uble vitamins are delivered to tisslles via chylomicrons; in the absence of chylomicron formation, the fat soluble
r,itamins will remarn in the intestinal epithelial ce1l, or not even be absorbed from the intestinal tract. Vitamin E is believed to be a major antioxidant factor in cells. AI1 of the other vitamins listed (B,, B, C, and niacin) are water-soluble vltamins, and do not require ch1'lomicron formatron for vitamin deliverY

in the mechanism of the phosphorolysls reaction ls still controversial, but it may play a role ln general acid-base catalysis. B, is not required for the PEP carboxykinase reaction, the
pyruvate carboxylase reaction (rvhich requires biotin), or the glucose-6-phosphatase reaction. Bu ls also not involved in regulating insulin secretion.

r: The answer is A: malic enzyme. The transcription of five key enzymes for fatty acid synthesrs are regulated by diet; malic enz)'me, acetyl-CoA carboxylase, citrate Iyase, fatty acld spLthase, and glucose-6-phosphate dehydrogenase. The other four enz)'rnes listed in the

,,f '

Theansweris D: B,r. AB,, deficiencywillblockthe methionine s)'nthase reaction, in whlch homocysteine reacts

with

N5-methyltetrahydrofoiate (THF)

to

regenerate

CH3- (CH2)n
Saturated

CH2

CH2

(CHz)*

,P
"iaaoO

+ C.2+ 2H*

fatty acyl CoA

Fatty acyl CoA desaturase

2 Cyt b5

(Fez

1- V

+\

/-t

2 Cyt b5

II

(FAD)

reductase .-/\

NADH + H*

Y
*o'-

2 Cyt b5

-/\ (Fe3*)

2 Cyt b5 reductase
(FADH2)

CH3- (CH2)n

CH = CH

(CHr).

- CtscoA

oo

Monosaturated

fatty acyl CoA

only Answer 4: Desaturation of fatty aclds in humans. The desaturase system requires a fatty acid which is at least 16 carbons 1ong, and can insert double bonds between carbons 4-5 , 5-6,6-7, and 9-10.

180

Chapter 20 methionine and free THE In the absence of such an actit' rt1,, the N5-methyl THF accumulates, and as that form is
the most stable form, eventually all foiate will be "trapped" as the N5-methyl derivative. N5-methyl THF cannot go

back to N5, N10-methl4ene THF; once the N5-methyl lorm rs synthesized, the folate is trapped in that fonn until the methionine sl,rrthase reaction occurs. Thus, folate ls still available, but in the wrong fom, the levels of N5,

N1O-methylene THF are too low to allow for th)rmidlne rynthesis, and the levels of N10-forrnyl THF are too 1ou to al1ow for sufficient purine syethesis Thus, a functional lolate deficlency occurs. Deficiencies in the other ritamins 1lsted (thiamine, niacin, nboflar'rn, and C) will not lead tc a functional folate deficiency The reaction involr'T ng folate and r,'itamin B,, is diagrammed belorv, along with other reactrons involved in homocysteine metabolism.

,,f
Homocysterne

+Dimethyl glycine

'Betaine

S-adenosyl homocysteine Adenosine

y'a -/\-a-crt'
Reaction pathu'ays that involr,e homocystetne. Delects rn numbered enzymes (1, methlonlne slnthase; 2, N5, Nl0-methylene-THF reduclase:

SAM

Cystathionine

_l ou Il'-''' Cysteine
The answer

d-Ketobutyrate. NH3

3, cystathioni.ne B-sl.nthase) lead to elevated homocysteine. A B,. deficiency u'111 block en4/rne 1, leading to a functional folate deficiency as all of the lolate wlil be trapped in the N5-methyl-THF form. This u.ill also lead to an accumulation of homocysteine in the blood.
methylene-THF reductase), and therefore reduce the amount of homocysteine converted to methionlne (reaction B of the figure). This would lead to a reduction of S-adenosylmethronine 1evels, and hypomethylation in the nervous system, which may lead to altered

is A: N5, N1O-methylenetetrahydrofolate

reductase.

thermolabile (temperature-sensitive)

N5,

NlO-methylene-THF reductase would reduce the amount of N5-methyl THF which can be formed (see rhe figure be1ow, reac[ion 7 utillzes the N5, N10-

Histidine
Sources of one-carbon groups

Serine Glycine

Ns,N1o-methylene FHo Donation oxidized groups

+ + \,
Ns-methyl

rrll,._lO l@ NAD(P)+ NAD(')H l.-'*r"

ii Ns{ormimino FHo

l@l

Tryptophan

HCOOH (formate)

Ns,Mo-methenyl FHa

l@
+

N1o-formyl FHo

of carbon

duMP --l TMp

-|,-NADH -)O -}\runo, r'[" e(

FHz -

l^
homocysteine

Donation of methyl group

NADPH --.,1

p4pp*

t''19r",n,onine

Adenosine S-adenosyl +L l7^Homocysteine -=---->

FHo

Purine biosynthesis

tR

l(s)

.1,

.l-

^6

#,.

,,

(sAM' meihYldonor)

1z s-adenosyt methionine -----l \

n-Cffs

The sources of carbon for the rerrahydrofolate pool are indicated by reactions 1-'1. The recipients of carbons from the THF pool are indicated by reactions 5-8. Reaction 7 is catallzed by the N5, N1O-methylenetetrahydrofolale reductase, while reacrion 8, which requlres vitamln B,r, is the methionine synthase reaction. Reaction 1 is catalyzed by the serine hydroxl.methyl transferase enzyrne.

Nutrition and Vrtamins 181

gene expression and a neural tube defect. Elevated homocystelne would also be evident. This is a common mutation in the general population, and can be overcome by taking folic acid. An inactivating mulalion in senne-hydroxymethyl transferase would reduce the major entry point of carbons into the THF pool, but there are other means to do thrs (reactions 2, 3, and 4 as indicated in the figure on page 180), so a loss of this enzyme would not result ln reduced S-adenosylmethlonine levels in the nen'ous system. A defect in ornithine transcarbamoylase (a urea cycle enzlnr-Le) would not aflect homocysteine and methionine metabolism. Similarly mutations in phenylalanrne hydroxylase, or tyrosine aminotransferase (enz1.mes involved in phenylalanlne and tyrosine metabollsm, respectlvely) would not affect homocysteine and methionine metabolism.

wlll not solve the problem, as methionine is lonnc .:

meai, and rs an essential amino acid. B,. suppler.nt:-tation also will not he1p, as the methionine synth.ts<
reaction, which requires B,r, is not altered; rl rs -1us. going at a reduced rate due to the lack of one ol rt-s substrates (N5-methyl THF). In thls condition, hom..cysteine accumulates due to the reduced effectiveness .. the methionine synthase reaction, so supplementaliLrll wrth homocysteine will not he1p. Iron supplemenialion wrll not help in this reaction sequence, as none ol the enzFnes are iron dependent.

The answer is C: Minimal insulin

release. The biochemi-

cai basis of the Atklns diet ls to minimize insuiin release.

insulin will promote glycogen and fat s)'nlhesis, and br minimizing its release (by eating a 1ow carbohldrate. high-protein diet), the ability of the liver to s).nthesize
these energy storage molecules

The answer is B: Folate supplementation. The mqor problem with the folic acld derivatives which are present in the patient is that almost all of the folate is trapped in the N5, NlO-methylene-THF form, leading to reduced Ievels of N5-methyl THTI and reduced regeneration of methionine from homocysteine. This leads to reduced S-adenosylmethionine ler.els, and hlpomethylation due to a lack of the methylating reagent. Supplementation with N5-methyl THF wiil overcome this block, and

will be greatly reduced.

(see

While amrno acids can stimulate insulin release

Table 20-1 and the figure below), the amount released is

about 10% that when glucose stimulates rnsuhn release.

A hrgh-protein diet wr1l not inhibit, allosterically, fatqacid slnthesis. A high-protein dlet does not increase g1ucagon release, nor does it reduce cortisol release. There wrli be increased urea production on such a diet, due to the lncreased ievel of protein degradation, but increasing urea production does not, by itself, lead to weight loss.

restore methionine levels to normal. Going meat free

120

J T

$
E

-J

roo
BO

High

protein
meal

c
C)

, ..--\
Nitrogen

o)

120

J ot
a ot or f

,/

I-6 tL
La

-=

rB0
E

g
o

140
Answer 9: PanelA indicates the increase in blood glucose, insulin, and glucagon levels after a high-carbohydrate meal. Note the scale lor lhe increase in in>ulin Ievels. Panel B indicates the release of lnsu1ln and glucagon in response [o a hlgh protein meal (note how 1itt1e insulin lncreases under these condtltons as compared to that in panel A). Also note how glucagon 1evels rncrease after the high-prorein meal, as compared to the high-carbohydraIe mea1.
0

o; orl -l or

eo-l

--.--v
I
Glucose

)' '= Lo
a

lnsulin

t= \
Lzo

-J tr

200
120

t- i Lrn

," a =
c

fl reo b
(g

E o)
o

110 100

e 160 c 3 t+o
= o
o
120
100

90

182

Chaprer 20

Table 20-1. Regulators of insulin release,

Effect

t0 The answer is D: Bu. AI1 three

neurotransmitlers

Major Regulators
Glucose

Minor Regulators
Amino acids
Neural input Gut hormonesb Epinephrine (adrenergic)
u+, stimulates bGut hormones that regulate fuel metabolism include, amongst many others, amylin, galanin, gastric inhibitory peptide, glucagon_ like peptide 1 and 2, neuropeptide y and somatostatin

of dihydroxyphenylalanlne, and histamine from

these reacLions lo occur.

(GABA, dopamine, and histamine) are derlved vja ciecar_ borl.lation of an amino acid precursor. Such amino acid decarboxlriation reactions require pyrldoxal phosphate (derived from Bu). GABA is denved from the decarbory._ lation of glutamate, dopamine from the decarboxylation
the

decarboxylation olhistidine. NAD, thiamine (B,), ribo_ flavin (Br), or cobalamin (B,r) are not required for any ol

1t

The answer is D: Tryptophan. This patient has the clas_ sic presentation of a carclnoid tumor. This type of tumor secretes serotonin which causes these classic s)rrnptoms.

The breakdown product of seroronin is 5-hydroxyindoleacetic acid (5-HIAA, see the figure below). Elevated

tT
H

tf

uH2

- CH- NHs

Nicotinamide moiety

\,,A*/
Tryptophan o, --'.L- gH,

n,o*J-t

-"G-JcH2-cN-riH3
H

gH

'

TryPtoPhan hYdroxYlase

5-Hydroxytryptophan

PLPFCO,

dopadecarboxytase

'"G--rcH2-cH2-r(H3
H

MAO-A

Serotonin

'"GJcH'-*-u
5-Hydroxyindoleacetaldehyde

f+

Acetyt CoA

f-t"oo.*

-*r.o
Macetyl Serotonin

[,- snrr,r f-tsnH

"rr"G7
H

o II cH2-cH2-NH-C-CH3

5-Hydroxyindole acetic acid

Melatonin

Answer I1: The biosy,rrthesis and degradarion of serotonin, leadlng to 5-HIAA (5-hydroxy indole
acetlc acld)

Nutrition and \.ritamrns f 83

ISmltters ia decar-

.ino acid
rrrsphate

:arboxy-

iation
the

levels of 5-HIAA in the urine conflrm a high 1eve1 of serotonin and the diagnosis of a carcinoid. Serotomn is derived from tryptophan, and the patient has a carcrnold tumor secreting serotonin. Elevated 1evels of a1anine, serine, tyroslne, or phenylalanine would not be observed in a patient with a carcinold tumor.
The answer is B: Easy bruising. The patient, due to the pancreatitis, is not able to adequately drgest triglycerides in the intestinal lumen, which is what leads to the ste-

t5

om

r. ribo,r any

The answer is A: Vitamin D. The child has r--.... which is due to a lack of vitamin D. Vitamin D,. -.-thesized via a circuitous route (see the figure be---'.'. and due to the parenis'(and childs) diet, there rs.:...ficrent vitamin D for the child to lorm healthr- bl:.. UV light is requlred to form the active form ol ri:,'.:: D, and the ch11d is also lacking exposure to sun _i" While, due to the diet, the child may become delicr:: for r,-itamin B,r, lack of B,, does not iead to these s\n:' toms. The symptoms are also not consistent u,rth a.;.-.. of vitamin K (whrch wouid lead to bleeding probler:rs

ol

he clas-

i tumor
rpioms.

rorpnier-ated

atorrhea. Fat-solub1e l.itamin absorptlon is dependent upon trlglyceride digestion and absorption, so under these condltions, the patient can become defi.crent in fat-soluble vitamins (A, D, E, and K). The loss of night vision is an early warning for lack of r.itamin A. If the patient is becoming deficient for r,T tamin K, brursrng would become a problem, due to rneffective clotting with slight internal damage. Nystagmus is a symptom of vitamin B, deficiency, a water-solubie vitamin. Dermatitis is a sl,mptom ol niacin defi.crency (vitamin Br). Loss of teeth can occur lvith a r.itamin C deficiency, and orange tonsils is not due to a vltamin deficiency, but rather [o a lack of ABC-I activity, and is indicatne of
Tangier disease.

folic acld (which would lead to anemia), or E (ltrss : protection against oxidative radicals). The pathri..t-'
active vitamin D formation is shown beiow.

CHs

r--r\ ./:-L-\-a
H"C

H-C-CH,-CH,-CH,-CH u.9 '"r.

,cH'

HoJt-J=.]J
7-Dehydrocholesierol

Skin {aru
t3
The answer is B: Triglycerides with medium-chain fatty

,,nn,

acids. Tiiglycerides containlng medium-chain fatty

acids are absorbed directly by the intestinai epithelial cells, and sent into the clrculation to the hver, bypassing the need to be incorporated lnto chylomicrons. Thus, conditions which might lead to fat malabsorptlon, such as pancreatic insufficiency, or decreased bile acid secretion, do not affect the absorption of these triglycerides. Long-chaln triglycerides do require the actions of pancreatic lipase and blle salts for absorptlon. Short-chain fatty acids are not utilized until they reach the large intestine, lvhere colonlc bacteria use them pnmari1y for energy. Thus, thelr nutrient value to humans is very low
The answer is E: Coconut oil. Coconut oil contains a high percentage (approximately 65olo) of medium-chain triglycerides. Butter also contains some medium-chain fatty acrds, but at a lower percentage than coconut oil (about 25o/o). Margarine is similar to butter in terms of length of fatty acids (although margarine contalns more unsaturated fatty acids and no cholesterol, as ii is not derived from an animal product). Olive oi1 is high in unsaturated fatty acids, with about T2obbetngoleic acid (an lS-carbon fatty acid with one double bond). Peanut oi1 is also high in unsaturated fatty acids but they are not medium-chain in length.
on of rdole

H-C-CH2-CH2-CH2-CH
cHs

?'. tf

,CHg

Cholecalciferol

Liver

+ 25-Hydroxycholecalcif erol KidneY I (9 PTH

Y

t^

-u-rydroxyiase

9H. -- cH" I zs1-"c H-C-CH2-CH2-CH2-C-OH

cHs

1,25-Dihydroxycholecalciferol
(1,25-(OH)2D3)
Sinthesis of active \-itamin D. 1, 25-di (OH)2 D3 is produced from 7-deh,.' drocholesterol, a precursor of cholesterol. In the skin, ultrariolet (L\-) Iig.: produces cholecalcilerol, lvhich is hydroxylated at the 25th position tn :.: lner and the 1st positlon in the kidney to form the actiye hormone.

L84 'i6 '

Chapter 20
The answer is E:

farln is

formation of y-carboxyglutamate. Wara vitamin K antagonist, and biocks the regeneration of actlve vj.tamin K after it has participated in irs reaction ol creating a y-carboxyglutamate residue (see below). in the absence of this side-chain modification, clotting proteins cannot bind to platelets, and the clot_ ting cascade is inhibited. Warfarin does not interfere

with platelet s)-nrhesis, nor does it alter phospholiprc bioqmthesis. Warfarln does not alter the transcriptioror translation of the clotting factors, and has no rela"tionship with vitamin E, which protects agarnst radica_ damage wtthln cel1s and tissues. As seen in the figure
be1ow,

warfarin blocks the actiyity of vltamin K epoxlde

reductase.

Vitamin
R

K.,

(Phylloquinone)

gH.
lt-

cHs

= -CHz-CH =C -CHzK2

(CH2-CH2

CH -CH2)3

Vitamin

(Menaquinone) cHs

R= -(CH2-CH=C-CHz)e-H
Vitamin
K3

(Menadione)

R=-H
B
CH,
I

\.r
I

CHz Prozymogen Carboxylated

zymogen

A:

Structures

of vltamin K

deriva-

CH.

t'

o*'oGlutamate residue
Vrtamin KH2 (hydroquinone)

c-H ./\ O=C C=O tt

tives. Phylloquinone is found in green ieaves, and rntestinal bacteria sl.nthe-

o-

o-

-Vitamin K

epoxide

y-Carboxyglutamate residue

oHo

od:"' \-^\4*
oH

size menaquinone. Humans convert menaquinone to a vrtamrn K active form. B: Vitamln K-dependent lormation of y-carboxyglutamate residues. The lrta, min K-dependent carboxylase uses a reduced form of vitamin K (KH.) as the
eleclron donor and converts r itamin K to an epoxide. Vitamin K epoxide is
reduced, in two steps, back to its actir.e lorm by the enzymes vitamin K epoxrde reductase and vitamin K reductase. As indicated on the frgure, warfann blocks the aclron of the vrtamrn K epoxide
reductase.

r RjHz t lL Il reductase =
uitu,in

-\

4-a/-n"^,^fi,n, ril#i:{
R1

^ \c io#:-'
^ARFAR,N
o

tv

vegetables. Green leafy vegetables contain large amounts of vitamin K, which wouid
The answer is C: Green leafy

{,8

overcome the effects of the warfarin. The other foods listed are poor sources of vitamin K (milk is fortified with vitamin D, orange-yellow vegetables are high in lrtamin A, trout, and beef are low in fat soluble vitamins, although organs, such as liver, would be high in vitamin K).

The answer is D: Formation of acetaldehyde adducts with cytoplasmic proteins. As shown on page 185, aceraldehyde, a product of ethanol metabolism, forms covalent

adducts with tubulin, which interferes with the movement and secretion of VLDL and other proteins normally secreted from the liver. It is not ethanol that is interfering with microtubule formation, but its metabolite that

r
il

Nutfltion and \
Hzo

itamns 185

Lipid
peroxidation

t.
Toxic

Amino

acids

-=+
i^

Binding to glutathione

..io
I

Proteins (clotting factors)

Proteins

NADPH
Ethanol

NADP+

NAD* NADH

/\:\

'

.-.

<

> AcetYlaldehYde

i 5J:": -injury (,

=#\*Acetvtatdehvde
Acetate

-'-

Acetylaldehyde

i ^ "f\v z NADH

w
Q)f (
VlOt_

of enzymes ALT AST

and -

(g

Glvcerol-3-P

\ --\
I

Swelling HzO

rlacylglycerols

---------------_>VlOr

I HrO/

nrot"in and lipid accumulation due to imPaired secretion

Answer 18: Alcohol-induced damage ro mlcrotubules, due to acetaldehyde accumuiation. Acetaldehyde-adduct formation decreases protein

Urr\hes\s anilrmpa\rsp'ro\r\\ srr\r\\o\, NrrrturNno\.r\n\t ilxsugt.Nrtrttu\'u\t Mnngt(xutttU\(t\(5t+Nutf rhe accumulation ol VLDL and protern wrthin the 1iver. Damage to the mitochondria slows the conversion of acetaldehyde to aceLate, thereb,v increasrng the availability of acetaldehyde to form adducts with a variety of liver proteins.

\tssiltlt$*S\t\tNt\

in[erferes with microtubule function. Ethanol does not directly rnhibit gene transcription (it actually leads to the transcription of MEOS, the microsomal ethanol oxidizing system). The levels of acetate formed are not sufficient to lower the pH of the liver cytosol. The ethanol carbons can be used for ketone body formation, but the formation of ketones is not related to the impaired secretion of VLDL. The toxic effects of ethanol on the
;h a-

liver are summarized in the flgure above.

nervous system, there is an indlcation that a strlct ketogenic diet can also help to alleviate the frequency of seizures in epileptrc patients. The ketogenic diet should be 80%, by weight, fat, wrth the other 20olo split between carbohydrates and protein If one can use fat containing medium-chain triglycerides, the diet appears to be more effective. The reason for the ketogenic diet having this effect has not yet been elucidated. Of the diets listed as choices, the one with the highest amount of fat would be the most ketogenic.

1l

19

t)' r-

The answer is A: 80% fat,201o combined carbohydrate and protein, by weight. The patient has epilepsy, a disorder of the nervous system which triggers, a[ limes, involuntary muscle movements (seizures). While there are drugs designed to reduce the electrical activity in the

20

The answer is C: Methylmalonic acid. The patient rs experienclng the symptoms of vitamin B,, deficiency The macrocytic anemia is due to a lack of precursors for DNA synthesis in the red blood ce11 precursors due to

186

Chapter 20

the B,, deficiency The numbness and tingling is due to hypomethylation in the nervous system, also due to the B, defrciency Vitamin B,, only participates in two reactions in humans. The first is the conversion of homocysteine to methionine, requiring N5-methyl THE The active form o[ B,, in that reaction is methyl-cobalamin (see the figure below). The second reaction is the conversion of L-methylmalonyl-CoA to succinyl-CoA. The active form of B,, in that reaction is adenosyl-cobalamin.

defi.ciency, neither of these reactions would proceed, so one would expect to see both homocysteine

In

8,,

and methylmalonic acld accumulate in the circulation. 8,, is not required for the metabollsm of acetic acid, ketone bodies, propionic acid (although propionic acid is converted to methylmalonyi-CoA, so in a B,, deficiency there may be a slight rise in propionic acid ievels as well), or succinate.

ASHB

CHz
I

CHz

H-C-NH" t" cooHomocysteine

Methylcobalamin

I+

iH COOl\l H-C-C-H / t t**-^'--'--.

( H c-scoA
lil
I

uretnyrrn'aronvicoa

l+

Adenosyl 1"12 cobalamin

'-1.
S
I

CHs,

.'

coot.-H-C]H | '-' H-C_H

C

CHz
I

ili

t_..*,-_.._. - SCoA,

l+ H-C-NH3 I cooMethionine

CHz

Succinyl CoA

-o*,-*---'

:,,

::

i'
;,,
t': i.:
i: i. it.

i,l

ii

i,l
1,.

i., : i..

i.
tl

i,
i.i

i. :l
!

i.

pfo-eine

Chapter

21

iion.
acrd,

;lcid

iefi:r'e1s

F{uman, Gem,eEics

and Cancer
This rhapter will test the rtader an basic concepts comcerning human genetits, and will also relate to the genetic aspects af cancer and specific signal transduction pathways in cryt*in cflnceYs"
A r,voman lr,ith ER*her2 breast cancer ce11s is lre3i..
u.ith an agent to block proliferatlon of such ce11s. Th,. agent r,vorks through which ol the following rval's? (A) Inhibiting DNA polymerase

QUESTIONS
Select the single best answer.

(B) Antagonrzing EGF-stimuiated ceil proliferation (C) Altenng estrogen's jnductron ol new gene traflsCnptlLr:(D) Stimulating the estrogen receptor to leat e the nucLe.l. (E) Blocking the synthesls of the estrogen receptor

A couple visits the obstetrician clue to a high lrequency ol miscarriages tn the family Suspecting a chromosomal abnorma1it1,, the physician orders karyotype analyses of the couple. For one of the tu'o, the follor'vrng karyotype was identifi.ed: '16, XX, derl5 t(9q;l5q). rhe primarl' reason lor the frequency of miscarriages is most 1lke1y which one of the follor,ving? (A) Potential regions of trisomy or monosom,v in the fertilized egg (B) Triplordy in the fertrlized egg (C) Chromosomal deletions in the ferttlized egg (D) Abnormal sex chromosomes in the fertilized egg (E) A Robertsonian translocation in the fertllized egg

A boy has a mild

case of ornithine 11xn562rf2per1.ls. de6ciencl', and has volunteere d to be part of a stud;' to tr' and controi the disease r''ia genetic engineering. A sr:rrabli

vector containing a functional ornithine lranscarbatroylase deficrency (OTC) gene has been developed, anc
needs to be targeted to which of the followrng organsi

(A) Bone marrow (B) Brain (C) Kidney (D) lntestine (E) Liver

A 5-year-o1d boy developed raprdly progressing muscle degeneration resuking in trouble standrng wlth observed
quadriceps, hamstring, and gluteal atrophy and hypertrophy of the calves. The chromosomal basis of this disease is most 1ikely which of the following? (A) A deletlon on the X chromosome (B) A translocation of the X chromosome with chromosome 12

4-year-old girl was brought to the Emergencr Department due to vomlting and lethargy. Blood work showed hyperammonemia and urinalysis demonstrated orotic aciduna. Neither of her parents hacl
ever expressed such symptoms. Genetically, this grrl is experiencing these symptoms due to u,hich one of the

following?

(C) Trisomy X
(D) A microdeletion on the Y chromosome

(E) A perlcentric inversion on the X chromosome

(A) Inherlting an autosomal recessive disorder (B) Inhenting an autosomal dominant disorder (C) Unequal X-inactivation during embryogenesis (D) Trisomy X gene dosage effects (E) Monosomy X gene dosage effects

r87

188 i6,

Chapter

2l
in the popularion. The female carrier frequency in the
population ls which of the followlng? (A) 1 in 5,000

A family had an i.nherited disease that spanned the last three gen"rations. A representatlon of a Southern blot of
the putative dlsease gene displayed the following:
GENERATION

123

(B) I in 10,000 (C) 1 in 20,000 (D) i in 50,000 (E) r in 100,000

10

The gene frequency for a rare autosomal recessrve dis.ur. i, I in 1,000 for the general population. The frequency o[ affected females is which of the following? (A) 1in 1,000,000 (B) 1 in 2,000,000

(C) I in 100,000 (D) 1 in 10,000 (E) 1 in 1,000

1l
The age of onset of the disease has decreased from the first to the thlrd generation. Thrs disorder may be due to which of the following genetic alterations?

A couple has had a child with a cleft lip and palate The relatrve risk of this couple having a second child with this malformation is best described as which of the following?

(A) Tianslocation (B) Triplet repeat expansion

(C) Trisomy

(D) Deletion

(A) Same as before their first chl1d was born (B) Same as the population in general (C) Greater than before their flrst child was born (D) Less than before thei.r first child was born (E) Less than the population ln general

(E)

Gene duplication

Considering the family ln the prevlous question, which of the following can best describe the finding of earlier age of onset of dtsease in each succeeding
generation?

l2

Chronlc myelogenous leukemia, due to the presence ol Philadelphla chromosome, leads to transformation due to the creati.on of a factor which is nonregulated and aberrant. Thls factor is best described as which of the follomng? (A) Tianscription factor (B) Growth factor recePtor (C) Growth factor

(A) Uniparental isodisomY (B) Anticipation (C) Malformation (D) Penetrance

(D) Tyroslne kinase

(E) Expressir'rtY
The disease frequency for an autosomai recessive dlsorder is one in a million. The carrier frequency for this a11ele is besl eslimated as which of the following?

(E) Ser/thr kinase

r3

A patient has been diagnosed with a melanoma,

and

(A) 1 in 500 (B) l in 1,000 (C) I in 2,000 (D) r in 50,000 (E) 1 in 1,000,000
A woman has a son with a rare X-linked recessive disease. which has a disease frequency of one in 10,000

molecu1a, analysis has indicated that the tumor has sustained a loss of pl6(INK4) activity (inhibltor of cyclin-dependent kinase 4). Such a gene would be best classified as which of the following?

(A) A dominant oncogene (B) A tumor suppressor (C) A proapoPtotic factor

(D) An antiapoptotic factor

(E) A growth factor

Human Genelics and

14

Cancer 189

Considering rhe pedigree shown below, rf 1-2 is a carrier of a rare autosomal recessive disease, what is the probabillty that IV-1 w111 have the disease?

t7

A family expresses an X-linked disease, and r'..1. --,.- . a marker available which can distinguish betr'. t.:2 polymorphic areas of the X chromosome, desrgn-.:- ^ as A and B. The family pedigree, along u.ith the pc morphic forms of the X chromosome lor each lan:member. is shown below. What is the probabilit;- th:: III-4 is a carrier of thrs disease?
.

il

(A) 1in4 (B) 1in8 (C) 1 in 16 (D) 1 rn 32 (E) I in 6'1 t5 In the

pedigree shown be1ow, assume 1-2 is a carrier for sickle cell disease, which occurs in the African Amerlcan community at a frequency of I in'100 live brrths. Assuming that all of the individuals in the pedigree are members of the African American communit),, what

Ut

IV

is the probabilrty that III-2 is a carrier for sickle cell

disease?

(A) 0% (B) 12 5% (C) 25olo (D) 50% (E) 100%

t8 Concerning the
mutation?

pedigree in the prer,-ious ques[lon, what ls the probability that indir.idual IV-1 is a carrier of the

(A) 0o/o (B) 12.5olo

(c)
ilt

25ok
50o/o

(D)

(E)
(A) (B)
).5o/o

100%

32.5ok

1S

(c)
(D)

37.5'.k
50o/o

(E)
l'6

1oo%

A 5-)rear-old presents to the pediatrician due to lethargy and tlredness. The red blood cel1 count is normal, but molecular analysis indicates that the chrld is producing 25ok ol the normal amount of o,-globin, compared to 100o/o of the normal amount of B-globin.
Further analysis shows a total deletron of the o-globin
genes from one chromosome. What might account for the lower than expected levels of o-globin belng
produced?

Utillzing the same pedigree as indicated rn the prer-ious question, what is the probability that III-l will have
sickle cell disease?

(A) 25ok (B) 12.5%

(c)
(D)

6.25ok 2.75ok
r .37 5ok

(E)

(A) (B) (C) (D) (E)

F-globin inhlbition of u-globin synthesis Enhanced expression of y-globin s).nthesis

Loss of a B-globin gene

Deletlon ol a third o,-globln gene Duplication of an o-globin gene

190 20

Chapter 21

A physlcian in a rural African clinic

sees a

child with

t*.ttirtg of rhe jaw, loosening of the teeth, and swollen

lymph nodes (see the picture below)' Karyotype analyof trtooa celis shows a translocalion betli"een chromosomes 8 and 14. This rapidly growing tumor is most llkeiy due to whrch of the follor'ving?

tit

(A) EBV activa[Ion (B) Bcr-abl activation (C) BC1-2 activation (D) Constitutive mYc exPression r E) ECF-rer'eplor acli\ ation

{\S\\TRS
I
lhe annrer is A Po'teltial rctoG in the tertiliz"d egg' -::: :::;--::e: 'i:r.s 3 iransiocaiion .l hLc:: --. :ltrl i i.--:e[s.'nian iransiocation, as those
od

trisory o. mono+omy

onlr' occur bets een acrocenlric chromosomes) betr','een chromosomes 9 and 1i. A piece of chromosome 9 (from the long ann, 9q) rs attached to the long arm o[ chromosome 15 (the derivatir.'e chromosome). As the carrier has all the genes present, she is normal. But when she makes gametes, the following combinations are possible: normal 9 and normal 15; normal 9 and long 15 (carry'rng a piece of 9q); shorter 9 (misslng the 9q area) and normal 15; and shorter 9 and long 15. When each of these four possibilities rs fertiltzed by a sperm carrylng a normal 9 and 15, the following four results are possible:

.: ::. a::r -.'!-l--::..:lllaIna:::la. -1 l-1--1:-aa.- -l:11-'----,r- --r' --- ' aaiLuiai r. ..... :- . -. -. ahrom,.r. n1c $;th -,:...'ilr: -:.r\:., :.llt s\-rnploms unless the ci-strophlr Sc:ll -' ':--. to these across the trvo chromosomes lan unirkeir e r e:- -: som)r X does nol lead to an\- s)Tnptom.. 11t. ,11'51roph-* gene is not on the Y chromosome. A periceninc Lnt e;slon within the X chromosome ls also unhkell'lo lead'..' dlsruption of the dystrophin gene.

-.:

(i)
(ii)

Normal 9 and 15 from mom, normal 9 and 15 from

dad-normal pregnancy and birth

Shorter 9 and long 15 from mom, normal 9 and 15 from dad-normal pregnancy and birth (this child will have the same translocatlon as the mom, and has all genes represented) Normal 9 and long 15 from mom, normal 9 and

(iii)

The answer is C: Altering estrogen's induction of new gene transcription. The patient is being given t2rmorrfen, which is a selective estrogen receptor modifler. -{s the woman's tumor cells are ER., the cel1s are expressing the estrogen receptor, and tamoxifen wili be effeetive in such cells. In breast cells, tamoxifen acts as an antagonist, blocking the actions of estrogen on the cells. In other tissues, however, [amoxifen acts as an agonlst, so the other tissues are responding normally to estrogen. Since the breasl cancer cells require a supply of estrogen to groq the use of tamoxilen wiii reduce the gror'vth rate of tumor cells. Tamoxifen does not stimulate the
estrogen receptor to leave the nucleus, nor does it block the synthesis of the estrogen receptor. Rather, the drug binds to the receptor and prevents estrogen from bind-

15 from dad-abnormal pregnancy, most likely leadlng to miscarriage. This embryo r'l'ill have trisomy 9q, and under most conditions, trisomy for a particular region of a chromosome is incompatible

ing to the receptor and altering gene transcriptlon

Tamoxifen does not inhibit DNA polymerase, nor does it antagonize epidermal growth factor (EGF)-stimulated cell proliferation, although in this tumor type (her2 ), there are no EGF receptors being expressed such that EGF would not have an effect on these cells.
The answer is E:

with live bi.rths. (iv) Shorter 9 and normal 15 from mom, normal 9 and 15 from dad-abnormal pregnancy, most iikely Ieading to miscarri.age. In this case, the embryo is monosomy for the 9q region, expressing too few genes for survival. The only monosomy that ieads
to
a

Liver.

The urea cycle occurs primarily

live birth is monosomY X.

These results are also summarized in Table 21-1

in the liver, so the defective gene only needs to be repaired in the liver for the cycle to become functional again. Targeting the vector to the other
ttssues

'2',

The answer is A: A deletion on the X chromosome. The

listed (bone marrow brarn, kidney, and intestine) will not resuit in a functional cycle, as those tissues do not
express the enzymes at a level sufficient for the cycie to proceed at an adequate rate.

boy is showing s).{nptoms of Duchenne muscular dystrophy, which is most often due to a deletion on the X Table 21-1. Father:9n and 15n
Father Gametes 9n 15n 9n 15n 9n 5n
'1

Mother:9n,9s, 15n, and

Mother Gametes
9n 15n 9n
151
'1

151

Genotype
9n9n 9n9n 9n9s 9n9s
'1

Outcome
Normal Trisomy 9q, Iethal event

5n'1 5n

15n151
1

9s 5n 9s
151

5n'1 5n

Monosomy 9q, lethal event

Normal, carrier of the translocation (same genotype as the mother)

9n 15n

15n151

lost piece of lVote: 9n and 1bn represent normal chromosomes 9 and 15.9s represents the chromosome I that has '151 a represents to chromosome 15.This chromosome 9 is missing a part of 9q. its long arm and that was translocated the lengthened chromosome 15, which is carrying a piece of 9q at its end'

L92 5

Chapter 21
The answer is C: Unequal l-inactivation during embryo' genesis. The girl is experiencing the symptoms of ornithine transcarbamoylase deficiency (OTC), whlch is a gene located on the x chromosome. Under usual conditions, women who are carriers of recessive mutated genes located on the X chromosome do not express symptoms of the disease. Hor,vever, due to gene dosage effects, during early embryogenesrs (the B to 16 ce11 stage of the embryo), one X chromosome is inaclivated in each cel1 (and becomes the Barr body) and remalns inactiYated in all luture daughter cells. What has happened in this child is unequal X-inactivation, in that the X chromosome carryng the nonmutated OTC gene was inactivated in the ma1orlty of primordlal cel1s, leadlng to the development of a liver in which the majority of ce11s expressed only the mutated form of OTC This led to a female having the

I in 500. This question requires an of Hard1,-Wsrnberg equilibrium for understanding population genetics ln which p' + 2pq, t 4' = | (p is the
The answer is A:

probability of having the normal allele, q 1s the probability of having the mutated a11ele lthus, p + 4 = I], qr is equal to the probability of having the disease, and 2pq represents the probability of berng a carrier for the
disease in the populatron). For this example, 42 = 10 so q = 10 3. Zpq, then, is 2 x 10 r, or one in 500 people
o,

will he a carrier for the

fhe answer is A:

disease.

in

5,000. In the case of an X-linked

sympioms of OTC deficiency Trisomy or monosomy X

will not lead to an OTC deficiency As the disease

gene is

X-linked, autosomal dominant and recessive inhentance patterns are not appropriate answer choices.

in 10,000 in this case) indicates that among 10,000 men, one would have the mutated gene on the X chromosome. Since women contain two X chromosomes, a collectron ol 5,000 women would represent 10,000 X chromosomes, and one of those X chromosomes would contain the mutation. This indicates that I in 5,000 women would be a carrier.
disease, the disease frequency (1

to
The answer is B: Triplet repeat

expansion. Tilplet repeat

in 2,000,000. Going back to the Hardy-Weinberg equilibriurn, q = ,g-'r (the gene freThe answer is B:

diseases (such as myotonic dystrophy or Fragile X slmdrome) are due to triplet repea[ expansions in or around a gene. The repeats tend to increase in number from one

quency of the mutated a11e1e), so that 42 = 10 6. Thus, the frequency o[ aflected indlviduals is one in a mi]lion.

generation to the next, which ls indicated in the Southern blot by larger-sized pieces of DNA hybridizing to the probe as the generations increase. As the triplet repeats increase in size, the disease usually becomes more severe, and the age of onset of symptoms is decreased. In some indivlduals with many repeats, no syrnptoms appear, and such i.ndir.iduals are consrdered "sleepers" and can pass the disorder on to their offspring. Nonaffected individuals also have a small number of repeats, but not enough to bring about disease. Tianslocalions, trisomy, deletions, or gene duplications would not show the pattern of signals seen in the Southern blot.
The answer is B: Anticipation. Anticipatron is the term used to describe a genetic dlsorder that increases ln sever-

However, the question asked for the frequency of affected females, which would be approximately one half of the aflected patients, leadrng to a lrequency of 1 in 2 million females would have the dlsorder, as would 1 in 2 million males (u,hich, when summed, gives an overall disease frequency of 2in2 milhon, or 1 in 1 mrllion indir.,iduals in the population would express the disorder).

lt

fhe answer is (:

ity from one generation to the next, as is often observed in triplet repeat disorders. Uniparental isodisomy ts
when a child inherits two copies of a chromosome from one parent. Malformatron is a birth defect due to environmental and genetic factors. Penetrance describes the percentage of people who develop symptoms upon lnheriting a genetlc disease (for example, inhentance o[ the BRCAI gene has a penetrance of 85% as 15% of the rvomen who lnherit the gene wili not develop breast
cancer). Expressir,rty describes the severity of symptoms an affected individual displays (individuals may show mrld or severe syrnptoms depending on the mutalion

Greater than before their first thild was Cleft 1ip and palate is a multifactorial disorder, requlring a large number of genes to interact in a way to create the condltion. Each parent needs to contribute a share of "altered" genes such that the condition is obsened. and this share needs to be above a threshold amount of "altered" genes. If the threshold is not realized, the condition ls not observed. Thus, there is a certain risk in the overall population for having a child with this condition. Once a couple has had a child with thls condition, we have identified two individuals who have a large number of "a1tered" genes. Thus, their risk, as compared to the risk of the population at large (the relative risk), is now greater due to the fact that the prior pregnancy indicated that this couple has a large number of "altered" genes between them.

born.

t2

which

rs

rnherited).

kinase. Philadelphla chromosome (a translocation of chromosomes 9 and 22) produces a new gene product, a fusion protern ofbcr and abl (bcr from chromosome 22 and abl from chromosome 9,
The answer is D: Tyrosine

Human Genetics and

C.',r:tr f 93
,

with the fusion protein being produced from the shorter chromosome 22). AbI is a tyrosine kinase, and when fused wlth bcr, it is constitutive and no longer properly regulated. The presence ol this unregulated kinase leads to a loss of ce1lu1ar growth control. The bcr-ab1
protein is not a transcription factor, growth factor receptor, growth factor, or ser/thr kinase. This transloca[ion is shown in the figure below.

13

The answer is B: A tumor

suppressor. CvcLn-;:: - - - .

kinase rnhibltors (CKl) act to block the aclirr;r .'- . ..---'-

that are activated by cyclins (see the figure :: When such an activity is lost (meaning that ::-. :rr-: products lrom both chromosomes are inactir-e '. *r- . -- - trolled cell proliferation can result. Since the ac..-....
must be lost, such genes are classrfied as tumor sLlpr:--:sors, as opposed to the dominant oncogenes, in uhi::an activity is gained via mutation or inappropriale ge:c regulation. The CKls are not invoh,ed in apopiosis. r-.. do they act as growth factors.

l)

922

C
l4

,@-c-abl
(-l-

u")
I

t l'=*.,

r1 ry\
/

I in 64. Since this is a rare autosomal recessive dlsorder, we can assume that the probabrlltr
The answer is E:

ol the individuals who married into the family

(11-i

I1-4) having the altered gene is zero. As such, the probabillty that 1l-2 or II-3 has inherited the mutated allele (and u'ill be a carrier) is 50% (a one in two chance ol getting the mutated allele from therr father, 1-2). The probability that 111-1 or 111-2 would inherlt the mutated al1e1e from their fathers rs also 50%. such that the overall probabilrty that I11-l and 111-2 would carry the

\
\

fr ^-/
@-bcr/abrrusionsene
Philadelphia chromosome

mutated allele is 50% times 50%, or 25olo. The probability that 1I1-1 would pass the mutated a11ele to 1\''-l is 50%, but since his probability of having the mutated a1lele in the first place is 25ok, the overall probabilit;of passing this gene is L2.5ob (1 in 8). This is also true for II1-2 passing the mutated allele to 1V-1. For 1V-1 to have the dlsease. both mutated alleles would have to be inherited, and the probabihty of that occurring

Growth iactor Receptor

--|>

lnitiates Ras/Ral signal pathway

+
lnduction of Activated CdK complexes

lnhibited complexes

@D --------r+
"*:. -------Answer 13: Control ol the G1/S transition in the cell cyc1e. The genes that encode cyclins and CDKs are oncogenes and the gene that encodes the
retinoblastoma protein (Rb) is a tumor-

>=(. ( )

.,1"\ u

\)

suppressor gene, as are the genes that encode CKIs (since the loss of therr actnity leads to tumor growth). CDK, cyclin-dependent kinase, CKI, cyclin-dependent kinase inhlbitor.

+
lncreased gene transcription Cell cycle progression

194

Chapter 21

is I2.5% times I2.5o/o, or

I in 64. These values

are

indicated in the pedigree below.

both events ro be rrue (the child to inherit rwo mutarecl alleles), the probabihrres need to be multiplied, and
0.275 times 0.051,ie1ds 0.0i375, or a j..375% chance.

IV
56% chance of having the disease (12.5% times 12,5%)

m 1.375%
(27.5% times S%)

The percentages in red indicate the probability of rhe individual belng a carner lor the disease, until you reach individual IV_1, in which case, the probabilitv is that of the person inheritlng the disease.

The percentages in red indicate the probability of the individual belng a carrier for the disease, except lor III_1, in which case. thev indicate rhe probability of thar individual having the disease.

17 15
The answer is B: 32.5%. For this problem, one cannot assume that the probability of a person marry.ing into the zero risk of carrlnng the sickle ..il Sir-r.. the disease frequencyis I in 400 (qr), the carrieifrequency ts 2pq, or I in 10. Thus, the probability that II_3 is a carrier ts 55ok (a one in rwo chance of inheriting the gene from
a

The answer is A:

0%.

Individual III_4 has inherited

A polyrorphic

one X chromosome from her father, which contains the

marker (indlcated by the red

family has

g*.

her morher, which is 50%, and a 1 in 20 chance of inher_ iring the gene from her father, which is 5olo Since either event can resuh in the child being a carrier, the probabili_ tles are added, yeelding 55olo). The probabiliry that III-2 wll be a carrier is rhe sum of the probabilities of inheriting
the mutated gene from either her morher (who has a 557o chance of being a carrier) or her father (who has a 10% chance of being a carrier). This comes out to 32.5ok (a 27.5o/o chance from mom and a 5ok chance from dad). These percentages are indicated in the pedigree below.

the figure below). This chromosome does not contain the disease mutatlon as her father does not express the dis_ ease. Her other X chromosome comes from her mother. and conrains the B pollmorphic marker. III_4,s brother has the disease, which came from his mother,s X chromosome with the A polprorphic marker (indicated in blue). Since 1il-4 did not inherit the mother,s X chromosome with the A pol;,moryhlc marker, she has no risk of belng a car_ ner for the disease. It is important to note in this question that there are two species of X chromosomes with the A polplorphic marker in thls family One carries the disease gene (from II-3), and the other does nor (from II_4. and also implied in I-1). This is indicated in the figure below

in

The percentages in red indicare the probability of the indlvidual

being a carrier lor the disease.

16

The answer is E: 1.375%. Based on rhe answer ro the last quesrion, it is known that the probabiiity of II-2 being a carrier is 55olo and of iI_1 being a carrier rs 10o/o. For III-I to have the disease, she must inherit the mutated al1eles from each parent. There is a 27.5ok chance of inheriting the mutated ailele from her mother and a 5ok chance of inheriting it from her father. For

*8

The answer is C: 251o. I-1 must be a carrier as her daughter (II-3) had a son wirh the disease. This means that one of the X chromosomes in I-1 carries the disease gene, although both X chromosomes display the A polymorphlc marker. lndividual II-2 has n j0% .hurr.. of inheritlng the disease gene with the A polymorphic marker from her mother (the X chromosome with the A polymorphic marker in II-2 had to come from her

.,.;.;::,i-

Human Genetrcs and mother, and there is a one in two chance that it rs the one wth the disease gene). Hower,er, based on the data in the pedigree, indir,rdual 1I-2 passed the X chromosome with the A polymorphic marker (indicated in red in the figure below), and a 50ak chance ol carrying the

Cancer

195

'

'

lII-2 (the other A marker X chromosome came from her father). 111-2 now has a 50o/o chance of passing the X chromosome, A polymorphic marker, and disease gene to her daughter IV-1. For IV-1 to be a carrler, all three events must occur, so the overall probabihty is 50% times 100% times 50o/o, or 25ab.
disease gene, to her daughter,

we are told that the patient is only producrng 25olo of the normal expected amount of u-globin protein. As there are sti1l two o,-globin genes remaining in the patient on chromosome 16, one possrbrlity is to have a deletion of one of the genes on that chromosome, ll.hrch u'ould reduce overall ct-globin gene expression to 25%. B-globin does not inhibit a-globin rynthesis, and enhanced expressron of y-globln wr11 not ailect u-globin expression. Duplica-

tion ol an u-globrn gene u.ould increase a-globin expression, whrch would decrease the seventy ol the disease. Similarly, deletion of a B-globin gene rvouid also alleviate the imbalance rn u-globin and B-globin syrthesis, and alleviate the severity ol the disorder.
20

The answer is D: onstitutive myc expression. The patient has Burkitt lymphoma, u'hich in 90% ol the
cases

is due to altered regulation of the myc gene (con-

stitulive activation ol transcription), due to a translocation of the myc gene such that it is controlled by an immunoglobulin promoter (r,hich is why this disorder results in abnormal blood cell proliferatron, as these are the cells that produce the immunoglobuIins). The translocation is shown ln the figure below.
Probabllity of being a carrier = 50% times '100% times 50%=

2s./.

lS

The answer is D: Deletion of a third cr-globin gene. Under normal condltions, a cell expresses 100o/o u-globrn protein. This comes from four o,-globin genes, which are transcribed equally (two copies of the o-globin gene on each

While Epsteln-Barr virus is thought to render individua1s susceptibie to Burkitt lymphoma, the oncogenic event 1s the misexpression of the myc gene. Bcr-ab1 is associated lvith chronic mvelogenous leukemia, Bci-2 overexpressron leads to a loss of apoptotic potentlal and is not associated with Burkitt lymphoma. EGFreceptor activation (similar to the erbB oncogene) also does not lead to these symptoms.

chromosome 16; see the figure below). Thus, each gene is contributing 25o/o of the total o-globin proteln in the ce11. When two of the genes are deleted, one would then expect to see a 50olo drop in total u-globin expression. However,

al\
l.a:-:...:::1

814

F'rl (}
f.'- -a

Chromosome 16

["']
(

HS40

tl'I

az

at

13

l \il il'l1 w.'

f
[,'
e'

f=-l

Chromosome E'

11

i]

LCRTGvAySP
Embryo: (zz = Gower
1

(zYz = Ponland
0"2E2= Qg\r\rq 2

Fetus:

c1212 cI2Y2

= HbF = HbF

Adult:

o'262=

A,

az9z= A

l(= lfr --/ \ \ fi-*

Ft't

@*',,*

'-

Genomrc organization of the globin genes. Note the two active coples of the o-globin gene on chromosome 16.

@\*vc oncogene