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BIO 310
Midterm 1 Study Guide
SBU
Summer 2013
N/A

MT 1 review questions:


1.What are the common features of all cells?
All Cells store their hereditary information as DNA. All cells
replicate their hereditary information by template
polymerization. Use RNA as an intermediary form of
hereditary information. Translate RNA into protein the way.
Use proteins as catalysts. USE ENERGY. Use the same
molecular building blocks. All cells are enclosed by plasma
membrane.
2.What are the basic differences between prokaryotic and
eukaryotic cells? Prokaryotic and Eukaryotic are distinguished
by their size, and type of organelles.
Prokaryotes are Bacteria, which arose 3.7 Billion years ago.
Eukaryotes include protists, animals, plants, fungi
Features held by Eukarotiytic CELLS not found in prokaryotes
Eukaryotes have a division of cells into nucleus and cytoplasm,
separated by nuclear envelope containing complex pore
structures. Eukaryotes have complex chromosomes composed
of DNA and associated proteins that are capable of compacting
into mitotic structures. Complex membranous cytoplasmic
organelles include ER Golgi COMPLEX, lysosomes,
endosomes,perpxisomes. Eukaryotes have specialized
cytoplasmic organelles for aerobic respiration and
photosynthesis. Eukaryotes have the ability to ingest
particulate material by enclosure withen plasma membrane
vesicles (phagocytosis). Eukaryotes undergo Cell division using

a Microtubule containing mitotic spindle that seperates

chromsomes. Eukaryotes have two copis of genes per cell
diploidy. Eukaryotes also have three different RNA
synthesizing enzymes(RNA polymerases).

3.What is the endosymbiotic theory of the origin of eukaryotic
cells?
Prokaryotes arose first and give rise to Eukaryotic cells. The
endosymbiont theory says that organelles in eukaryotic cells
(mitochondria and chloroplasts) evolved from smaller
prokaryotic cells

4.What are some mechanisms of gene divergence?
Example 1. Divergence of originally identical genes from
different mutations in sister lineages
Ortholog Genes: Genes in different species that share a
common ancestor, they differ gradually, probably conserve
function
Example 2. Gene duplication and divergence
Paralog: Genes that arise by duplication and divergence and
evolve to have and evolve to have different functions
Example 3. Lateral Gene Transfer
More common in Prokaryotes Conjugation, Transformation,
Transduction, Symbiosis


5.)Explain the features of the last common ancestor
Common ancestor of all modern cells existed 3.5 bya had about
600 genes. Life diverged from this common origin by natural
selection of favorable mutations. Lateral Gene Transfer among
species has contributed to diversity.


6.)Explain the evolutionary relationship between eukaryotes,
archaea, and bacteria. We resemble Archaea in how we handle
genetic information (Replication,Transciption,Translation). We
resemble bacteria in Metabolism and Energy Conversion.
Archaea are the prokaryotes that often but not always live in
unsual enviroments or Extremephilles. Methanogens convert
CO2 and H2 into Methane. Halophilles: Dead Sea or Brine Pools
with salinity 5M.Acidophilles:ph0 ababoned mine shafts.
Hyperphilles: 121C.More than 200 gene families are common
to all 3 branches of life. Presence of all life Eukaryotes and
Prokaryotes.
7.) What is the role of hydrogen bonds in protein structure? in
DNA structure? in RNA structure?
In protein Structure the Hydrogen Bonds help make the
secondary structure. The Coils and Folds of Secondary
structure results from H-Bonds between the repeating
constituents of the polypeptide backbone. C=0 H-N. Typical
Secondary Structures are a coil called an alpha helix and a
folded structure called a beta-pleated sheet.3.6 Residues per
turn of Right Handed helix places Carbonyl oxygen of residue 1
with amide proton of residue 5. Hydrogen Bonds also
contribute to Tertiary Structures of Proteins. In Beta Sheets

Amide protons and Carbonyl Oxygen hydrogen bond with

neighboring strands .
In DNA Structure there is Hydrogen Bonds between Base pairs
of complementary strands of DNA.
In RNA Structure .RNA Tertiary Structures are Reminiscent to
those found on Proteins. 3 structures involve 2 structures
interacting with one another. The 2OH on ribose can HBond
with other bases. These 3 structures can have enzymatic
activities structural and regulatory activities. RNA has this
extra 2 Hydroxyl Group. That Can H Bond with other bases.
8. What forces stabilize correctly folded proteins?
In Folded proteins the Hydrophobic Residues are located
primarily in the middle or center of the protein, particularly in
the vicinity of the central Heme group. The Hydrophilic Side
chains, which are showing, are located primarily at the surface
of the protein where they contact surrounding aqueous
medium. So Alpha helical Coiled Coil stabilize by

hydrophobic interactions

9.Explain the advantages and disadvantages of light and

electron microscopy. Know the limits of magnification and
resolution for each.
Cells and Subcellular Structures cannot be seen without a
Microscope. In Light Microscope visible light passes through a
specimen and then through light lenses, which magnify the
image Compound: Product of Ocular and Objective. Light
Microscopes can magnify effectively to about 1000 times the
size the actual specimen. The Minimum resolution of an Light
microscope is about 200 nanometers because of the
wavelength of a beam of light. Ocular=10x Objective =100x

40=Minimum Magnification
1000=Max Magnification
Fluorescenece =Absorption of a photon raises an electron to an
excited state. A longer wavelength photon is emitted when the
electron falls back to the ground state.
Electron Microscopy: Electron Microscopes use Magnets to
focus electron beams similar to how a light microscope uses
lenses to focus light beams. The electrons are then directed to
photographic film to produce an image. Because electron
beams have much shorter wavelengths they have 1000 times
the resolving power of a light beam .2nm 1million times the
Human eye.
LIGHT Microscope: 1000 Magnification, 200 nm Resolution
Electron Microscope: 1000,000 magnifications .2nm Resolution
Most Subcellular Structures or organelles are too small to be
resolved by a Light Microscope.
Two Basic Types of Electron Microscopes are used to study
subcellular structures
Scanning Electron Microscope: Focus a beam of electrons
onto the surface of a specimen providing images that look 3D
Transmission Electron Microscope: Focus a beam of
electrons through thin sections of a specimen Used to study the
internal ultrastructure of cells.
Freeze Fracture Microscopy: Cells are frozen and then a knife
is used to crack them open. The Crack often passes through the
interior of plasma and internal membranes. The Bumps that
appear are usually large proteins embedded in the interior of
the membrane.


10.) Describe methods to purify proteins.
Proteins can be separated by column Chromatography.
Purification is the Stepwise removal of Contaminants using
successive techniques that take advantage of different
properties of proteins such as size, charge, shape.
There is Gel Filtration Chromatography, Ion Exchange
Chromatography, Affinity Chromatography.
Gel Filtration Chromatography: Separates proteins by size
using tiny porous beads. Bigger proteins will come out First!!!
Dont interact with the porous bead. Smaller Molecules are
retarded in the beads SLOWED DOWN!! Also called Size
exclusion.
Ion-Exchange Chromatography: The overall charge of a
protein is the summation of all charges of the individual amino
acids which in turns depends on the ph of the solution
Anion Exchange Columns: + Beads DEAEC cellulose. + charges
and neutral charges will come out first
Cation Exchange Columns: - Beads CM Cellulose. - charges will
come out first.
By adjusting the ionic strength and ph of the solution, proteins
can be bound and eluted.
Isoelectric Point pI: the ph at which the protein carries NO
net charge.
Proteins have a net negative charge at pH above their pI and a
positive charge at pH below their pI. By adjusting the ionic
strength and pH of the solution, proteins can be bound and
then eluted based on their isoelectric point. For example a

protein will bind to negatively charged beads at pH below their

pI but if you RAISE the pH the protein will become neutral or
negatively charged and it will elute.
Proteins can be separated by affinity. Affinity Chromatography
exploits specific biological interactions between molecules.
Enzyme-Substrate or Antibody-Antigen or Ligand-Receptor
Also can use DNA sequences bound to beads to recover specific
DNA binding proteins. Best way of purifying any protein.
Affinity Tags(Epitope tags) for purification and immune-
localization. Fusing protein to a known tag. Epitope tag can
easily be recognize by another protein. Gene of interest and
express it in Bacteria in a way that has a tag coupled to it. Use
that hook to purify it ways of cleaving that tag.
Gel Electrophoresis: Can also be used to separate proteins. But
proteins need to be coated with charge first. SDS Page.
11.)Explain how DNA sequencing works.
DNA can be sequence by dideoxy chain terminators
dideoxyRibonucleoside Triphosphate prevents strand
extension
at
3
end.
Rare
incorporation
of
dideoxyribonucleoside by DNA polymerase blocks further
growth of the DNA molecule. DNA sequencing by chain
termination with di-deoxynucleotides. A Blast Search can be
used to find similar amino acid sequences in varitety of
organisms.
Micro-Arrays can be used to assess genome wide changes in
gene expression or copy number.
Gel Electrophoresis: Separate DNA based on size . DNA(-)
charge runs to positive field.
Mutagenesis: Change a DNA that encodes a protein see if
affected phenotype of a cell or phenotype of organism.

STRAIGHT FORWARD.
Genetic Modification: Use Recombination to replace wild-type
gene with Mutant gene. Gene Replacement :only mutant gene
is active Gene Knockout: No active gene is present Gene
Addition: Both normal and mutant genes are active.
Mutations in DNA can affect their protein product in different
ways.
We can use a Yeast two Hybrid Screen for Protein-Protein
interactions
Cells that interact with protein of interest an indicator light
goes on. In most cases to turn on the expression of a gene you
need a transcription factor sits on Regulatory region of a genes
so it Recognizes a particular DNA sequence when its there it
recruits RNA polymerase and all other Transcription factors to
come and express that gene. There is a DNA Binding domain
that recognizes the promoter region, then trans- activating
domain that summarizes RNA polymerases and other
transcription factors. So when DNA-Binding Domain and Trans
activating Domain and if these two come together they activate
Target proteins .So what makes proteins of interest interact?
What makes them come together is if one of them is attached
to one protein and the other attached to another protein and
these two proteins interact in Real life.
DNA sequencing
deoxynucleotides

by

chain-termination

with

di-

12.)Explain how PCR works.
PCR can be used to obtain genomic or cDNA clones. You
amplify gene!!!! The polymerase chain reaction uses a heat-

stable DNA polymerase and two primers (oligonucleotides,

each complementary to one of the ends of a DNA sequence of
interest) to synthesize a strand of DNA complementary to
another DNA STRAND. This reaction is repeated to double the
number of copies.
Double Stranded DNA with sequence of interest is denatured
by heating to separate the two strands. An excess of
oligonucleotide primers complementary to the ends of the
sequence of interest are added and allowed to bind by base
pairing. DNA polymerase synthesizes complementary strands
starting from the primers This CYCLE is repeated many times
to amplify the sequence of interest.
Heat for a minute this denatures the DNA sequence of interest
you then add complementary oligonucleotide primers, cool for
annealing. Synthesize complementary strands. This doubles
the numbers of identical DNA. .
Micro-Arrays: Display thousands of tiny spots on a glass slide,
each with a particular DNA SEQUENCE or protein. This allows
many reactions to be monitored in parallel.


13.)What is the fluid mosaic model of membrane structure?
The Basic structure of the Membrane is a lipid Bilayer with two
sheets of phospholipids arranged tail to tail. Proteins are
dispersed throughout the membrane in a mosaic pattern.
Proteins contribute to membrane structure and function. Some
proteins span the membrane while others are confined to
inner and outer leaflets. The membrane is fluid: Most proteins
and lipids can move freely withen each of the two leaflets
.Although they cannot move easily between the two leaflets.

The lipid bilayer serves as a Hydrophobic barrier, confining

Hydrophillic molecules to the inside or the outside of the cell.
The Lipids are also involved in regulatory events.
14). Explain some lines of evidence that supports the fluid
mosaic model of membrane structure?
We can visualize Membrane Proteins by Freeze Fracture
Microscopy, we freeze the cell, we then fracture the frozen cell
wih a knife which splits the Lipid Bilayer in half when then
view this under a scanning Electron Microscope and see that
proteins are actually spanning the membrane. We can also
visualize the lateral Movement of Lipids using Fluoresnce
Photo bleaching in which we see Lipid Movement in the
membrane because the Fluorence label comes back from
Unbleached parts. Color is recovered very quickly suggest
Lipid Molecules are moving very quickly . Lipid Movement
1micrometer per second. We can also visualize protein
movement but proteins move at a much slower rate. We label
two different proteins and watch how they move after
incubation. Not all proteins move at the same RATE
15) What exactly is meant by Fluidity ?
Fluidity refers to the Viscosity of the Lipid Bilayer of a cell
membrane; Fluidity depends on the Lipid Composition.
Saturated Fats make membrane less fluid. Unsaturated fats
make membrane more fluid. Lipids have rapid lateral
movement.
16.)Understand the basic structure of lipids: fatty acids, polar
head groups, and cholesterol.
Membrane Lipids:
Cholesterol

Phospholipids,

Sphingolipids,

Phospholipids: Also called Glycerophospholipids

and

Are diacylglycerides with small functional head groups linked

to the glycerol backbone by phosphate ester bonds.
Polor Head Groups:Ethanolamine Choline Serine Glycerol
Inositol Inositol 45 biphosphate
Ethanolamine and Choline neutralize the negatively charged
phosphate Phospholipid is then Neutral.
Serine had one positive and one negative charged and the rest
is uncharged so the phospholipid has a negative charge.
Sphingosine is equivalent to glycerol and one Fatty Acid. A 2nd
fatty acid is attached by an amide bond to carbon 2.
Glycosphingolipids has head group that is sugar
Cholesterol is key component of the plasma membrane it
helps make it a little bit solid, and helps make it less
permeable. Cholesterol is involved in the synthesis of steroid
Hormones such as estradiol, testerone,cortisol also Vitamin D.
Cholesterol helps regulate fluidity, the presence of cholesterol
is a major determinant of membrane structure, function and
activity. Polor Head groups of membrane are stabilize by
interaction with water. The Hydrophobic portions are stabilize
by Hydrophobic interactions. (Fatty Acids toward middle )
Model of Phosphatidylcholine Bilayer: Excluding water
raises entropy, makes structure thermodynamically favorable.
Sphingomyelin and Cholesterol are together on the outer
leaflet. Plasma Membrane has much more Cholesterol and
Sphinomyelin then internal membranes. The presence of
Cholesterol makes the Plasma membrane less permeable
,while maintaining fluidity . The membrane is asymmetrical :PS
and PI , with net negative charge , face the cytoplasm, making
the cytoplasmic side negatively charged.

Lipid Rafts: Are enriched in SM, GS,and cholesterol

(Microdomains different kinds of molecules. )
Most Membrane Lipids are synthesized entirely in the ER.
Enzymes called Flippases can flip the phospholipid to the
opposite leaflet.

17.)What is the difference between an integral protein and a
peripheral protein?
Integral Proteins: May be removed by the membrane by the
use of strong detergents, organic solvents or protein
denaturation They are CRITICAL to the Structural integrity of
the membrane, and constitute about 70% of membrane
proteins. When removed, they sometimes still have lipids
attached. Most proteins are integral membrane proteins.
Pepherial Proteins: Are located outside the Bilayer, and are
associated with the membrane by noncovalent bonds. They
may be associated with lipids or proteins. They may be
associated with lipids or proteins. They are removed by mild
treatments :aqueous buffers or changes in the ionic strength of
cells. They are free of Lipids when removed.

18. Explain the features of integral membrane proteins.
Typically Trans membrane domain are alpha helical. Integral
Membrane Proteins are typically alpha helical. Intergral
Membrane Proteins can have a single alpha helix with lipid.
Multiple helices, rolled-up beta barrel. Attached by
Hydrophobic domain, prenylated attachement. GPI anchored
and Transmembrane integral protein.

In Hydropathy plot , if there is lots of Hydrophobic Amino

Acids this might be indicative of a trans membrane domain.
Glycophorin A is an integral membrane protein with a single
transmembrane domain Gly-X-X-X-Gly.
Hydrophobicity is measured by the free energy required to
transfer each segment of the polypeptide from a Nonpolor
solvent to an aqueous medium. Peaks above the o line are
interpreted as a trans membrane domain. In Intergral
Membrane proteins alpha-helical H Bonding increases
Hydrophobicity. Very difficult to study integral membrane
proteins.

19.) List the ways that peripheral proteins can be associated
with membranes.
Six modes of association of peripheral proteins with lipid
bilayers. Peripheral Proteins can be associated because they
have some small fatty acid. Six modes are acyl
chain,electrostatic,partial insertion, or bind directly to integral
proteins.
C-terminal Isoprenoid
N-terminal Myristoyl
C-terminal GPI
Electrostatic interaction between calcium binding proteins
bind to PS.
Hydrophobic Helices on enzyme pG synthase partially
penetrate bilayer. Peripheral protein associates with Trans
membrane protein.

A membrane skeleton stabilizes the cytoplasmic surface of red

blood cell plasma membranes. Inherited deficiencies of these
proteins cause fragile red blood cells and hemolytic anemia.


20.Are membranes symmetrical or asymmetrical? How is this
regulated? The membrane is asymmetrical PS& PI with net
negative charge face the cytoplasm making the cytoplasmic
side negatively charged. Cholesterol help pack the membrane a
little bit tight. Flippases help regulate asymmetry by flipping
phospholipids to the opposite leaflet. Membrane asymmetry
different composition inside and out or fixed orientation of
proteins spanning the membrane. The membrane asymmetry
allows the cell to automatically differ its intracellular
environment from that of the excisting extracellular
environment.

21.Explain the common features and differences between
uniporters, antiporters, symporters?
Uniport: One single substrate down its concentration gradient.
Antiports: (Exchangers) Driving Ion in one direction, driven
substrate in other.
Symporters(Co transporters) Driving ion and substrate both
in same direction
Antiport and Symport are both examples of coupled transport.
In animal Cells, the driving Ion is usually Na sodium.
All of these are membrane carriers the concentration gradient

determines the movement. Carrier is not determining direction

molecule is moving. Carriers are membrane proteins that
alternate between 2 conformations that bind and transport
solutes across membranes, down concentration gradients. The
conformational change is the result of substrate binding. The
direction is determined by the substrate concentration on
either side. Substrate can bind on either side when it binds
cause conformational change.




22.)What are the energy sources used by pumps, carriers, and
channels to transport substances across membranes?
Membrane Pumps(primary Transporters) use energy from
ATP hydrolysis, light absorption or redox reactions to move
ions or other solutes up concentration gradients
Three Types of Tranport Atpases
F-type Atpase: Use either Atp Hydroylsis to transport protons
up concentration gradient or proton gradient to synthesize
Atp.
P-type Atpase
ABC Transporter
Membrane Channals: Allow Ions to cross a membrane down
their electrochemical gradient .
Membrane Channels cooperate with pumps, carriers to

transport water and Ions regulate electrical potential across

membranes signaling.
Membrane Carriers: The energy source of Membrane carriers is
the concentration gradient of the substance being transported.
Also carriers are membrane proteins that alternate between 2
conformations that bind and transport solutes across
membranes ,DOWN concentration gradients.


23.)Why does the rate of transport by a carrier plateau?
Rate of Transport reaches a maximum because the transporter
is saturated. Transport of Radioactive glucose into Red Blood
Cells established the existence of Membrane Carriers.
Transporter mediated diffusion has an accelerated initial rate
of entry. It also reached a Maximum when transporter is
saturated.
Experimental Evidence for the existence of Symporters
Extracellular Na is required for glucose uptake by SGLT 1
Gatorade takes advantage that Glucose and sodium use the
same molecule of transport to get inside Intestinal Cells and
eventually into the Bloodstream. The SGLT transporter
oscillates betweeb 2 states. The binding of Na and glucose is
Cooperative the binding of one increases the affinity for the
other. The conformational change occurs when only both or
neither ligand binds. Also GLUT 5 glucose uniporter transport
glucose down its gradient from epithelial cells to bloodstream.

24.)How does a K-channel discriminate between K and other

ions?
K channel has 2 transmembrane helices and a Ploop has
binding sites for 4k ions. The Voltage-gated potassium channel
contains six membrane spanning helices HAS A PENTA
PEPTIDE. A single channel has four subunits arranged to creat
an ion-conducting pore the channel can be opened closed or
inactivated. Cystral Structure shows that a bacterial K channel
shows a short AMINO ACID Sequence that selects for K and no
other Ions. Only potassium that is not hydrated can fit in the
potassium channel. IF YOU ALTER ANY AMINO ACIDS IT
DOESNT WORK. Penta peptide negatively charged Carbonyl
forms a pore that is just the rite size to accomadate a strip of
potassium ion that is not hydrated!!!!!!!!!!!
25.How does aquaporin block the transport of protons?
Aquaporin Channels are specialized protein channels that
allow passive movement of water. The asparagines in the
Aquaporin channel teher a central water molecule, which can
no longer Hydrogen bond, preventing the flow of protons
through the aquaporin channel. Asparagines in Aquaporin
channels prevent the hoping of protons.
The placement of Aquaporin Channels in the Renal Collecting
Duct is under the control of Antidiuretic Hormone Vasopressin.

26.)What substrates are transported by the 3 families of
transport ATPases?
F-type ATPASE: Mainly H Use either ATP Hydrolysis to
transport protons UP concentration gradient or a proton
gradient to synthesize ATP.

P-TYPE ATPASE: Named P-TYPE BECAUSE THEY BECOME

TRANSIENTLY PHOSPHORYLATED Use by all forms of life to
Pump cations across membranes IN ANIMAL CELLS NA K
Atpase forms gradient, pumping Na out K in both UP THEIR
CONCENTRATION GRADIENT. In all Eukaryotes a Ca ATPASE
in the plasma membrane and in the endoplasmic reticulum,
clears Ca from the cytoplasm. Ca bind to the TM helices ATP
binds to the N domain. Conformational CHANGE reduces
affinity for Ca,pumps CA Into lumen and promotes
dephoshorylation of D351.
ABC Transporter: Mean ATP Binding cassette. These are
found in all proks and euks. They are modular in design, with
TM domains and cytoplasmic ATPASE/regulatory domains.
Transport a Variety of Substances. Flippase is an example of
ABC Transporter. In Eukaryotes, almost all ABC
TRANSPORTER export substances from cytosol to extracellular
space or to a membrane bound intracellular compartment such
as ER OR MITOCHONDRIA. In Bacteria help take things INTO
the cell Transport to Cytosol. ABC Transporters Variation of a
Common theme ATP Binding induces a conformational change
that exposes substrate-binding pocket to the opposite face.
Multiple Drug Resistance of Human
overexpression of MDR ABC Transporter.

Tumors

from

MDR ABC Transporter: Found in plasma membrane of all

cells does a good job of pumping out anything from the cell that
doesnt belong there. Upregulation of MDR ,YOU THEN NEED
HIGH DOASGE OF DRUG.


27.)Which transport ATPase family uses a covalent,

phosphorylated protein intermediate?

P-type Atpase :Use a High energy covalent B aspartyl
phosphate intermediate
Ca ATPASE : Pumps Ca out of the Cytoplasm into the
Endoplamic reticulum ATP Hydrolysis provides energy to
move Ca across the membrane up a steep concentration
gradient.




28.)How do transport ATPases use ATP hydrolysis to transport
substances across membranes? Use energy from ATP
Hydrolysis to move ions or other solutes up concentration
gradients.


29. Why is the size of a genome not directly proportional to the
number of genes? The genome is not Homogenous in structure
and function and the structural features of the chromosome
are a consequence of the genome itself. Also the Human
Genome has a lower gene density and a greater proportion of
non-coding and Repetitive DNA. Less then 2 % of the genome
actually encodes proteins , there is huge vast stretch of DNA
BETWEEN each gene we have a much lower gene density then
simpler organisms .



30.What is meant by gene density?
The amount of genes per particular DNA. Human genome is
much less dense there is much fewer genes per particular
length of genome. Our genome is fairly poor in genes. LESS
then 2% of the genome actually encodes proteins. We have
much lower gene density then simpler organisms.
31.What is the structure of a chromosome?
Principle features of Eukaryotic chromosomes include
centromeres and telomeres. Has 4 telomeres, A p and q arm.
Sister chromatid there is two per chromosome.


32.)Explain the structure and function of centromeres.
Centromere is the region of the Chromosome that interacts
with proteins to form Kinechores . Centromere itself is full of
repetitive DNA. There are no genes in the Centromere.
Centromere is place where Kinechore forms. Centromere has a
Unique structure only place where Mitotic spindle attaches,
and pull replicating chromosomes to opposite directions
33.)What is the difference between a centromere and a
kinetochore?
Kinechore: Complex disk of proteins. Proteins in Kinechore
need to be recognize by Mitotic Spindle. Centromere is usually
in the middle of the chromosome. Centromere is place
Kinechore forms. Centromere is a unique structure only place

where Mitotic Spindle Attaches proteins in the Kinechore

forms. THE Kinetochore is a protein complex that connects the
Centromere to the Mitotic Spindle. The Kinechore contains a
huge number of factors regulatory proteins.
34.)Explain why replication of the ends of chromosomes
presents a challenge and how telomerase solves this problem.
The end replication problem limitations of DNA Polymerase
creates problems for the linear DNA of eukaryotic
chromosomes. The usual Replication machinery provides no
way to complete the 5 ENDS, so repeated rounds of replication
produce shorter DNA Molecules. In order to Replicate, very end
of Molecule you would need a primer out in space. There is no
guarantee that the primer is going to end at the very last
nucleotide. You are going to lose end chromosomes every time
DNA replicates. Ends of chromosomes dont encode proteins.
Telomeres are repeated hexamers six nucleotides 4
telomeres per chromosome. So you dont have to worry about
losing genetic information As telomeres shorten they reach
end of mitotic lifetimes they dont die this is phenotype of
aging. Cells stop dividing because their telomeres have gotten
short.
Telomerase repairs shorten telomeres and give cells
immortality. Telomerase is composed of a protein with
Reverse Transcriptase activity and an associated RNA .
Telomerase replicates chromosome ends !!!!


35.What is the structure and function of histone proteins?

The proteins that DNA is associated are called Histones.

Histones are a family of small Basic proteins that can be
associated with DNA. Histones are rich in Amino acids that
amines in their R group like Lysine and Argine positive charge
of these R groups is attracted to negatively charged DNA . If
you want to compact DNA makes senses to wrap around
positively charged proteins. Histone fold is called a Z fold.
Histones are a family of small basic proteins C-terminal end of
Histone proteins allow them to dimerize . Histone proteins
associate with one another they also associate with DNA . N-
terminal Histones are in a less saturated 3 dimensional shape.
Extend out of the Nucleosome and play a major role in
chromatin structure.

36.What is a nucleosome, and what is its structure?
The first level of folding involves coiling DNA around a protein
core to yield a Nucleosome. This shortens DNA about sevenfold
relative to naked DNA. Each core Histone has a compact
domain of 70 to 100 amino acid residues that adopt a
characteristic Z shaped Histone fold consisiting of a long a-
helix flanked by two shorter a helices.
Nucleosome is the Basic Unit of DNA Packaging (8 globular
proteins in the middle consisting of a segment of DNA wound
in a sequence around eight Histone cores. Nucleosome
structure protects the DNA from Digestion. All DNA is in the
Nuclesomes
37. What is the difference between heterochromatin and
euchromatin?
Telomeres

and

Centromeres

are

refered

to

as

Heterochromatin. Genes with Highly packed Heterochromatin

are usually not expressed
Euchromatin contains almost all the genes both actively
transcribed and quiescent.
Heterochromatin: Is transcriptionally inert and is generally
more condensed then euchromatin it was initially recognize
because it stains more darkly with DNA binding dyes than the
remainder of the nucleus.
All DNA is in the Nucleosomes but beyond the level of folding
and compacting differ whether talking about Heterochromatin
or Euchromatin. Euchromatin is read by Transcription
Machinary.
38.How does the spatial organization of chromatin within the
nucleus correlate with gene expression? A typical Nucleus has
both euchromatin and Heterochromatin, the latter usually
being concentrated near the nuclear envelope and around the
nucleoli. Much of the interior of nuclei is occupied by pale
staining euchromatin rich in actively transcribing genes.
Heterochromatin might also be pointing to Nuclear Lamina.


39. Explain epigenomics and how specific covalent
modifications of histones and DNA regulate the state of
compaction of chromosomes and their availability for
transcription.
Process that instruct our cells how and when to read the DNA
BLUEPRINT.
So Histone Tails protrude from the Nucleosome Core.

Modifications to both Histone Tails and DNA are indicative if

DNA is active or not .
Constititutive Heterochromatin: Is associated with special
types of DNA sequences such as satellite DNA that are
packaged into a particular closed conformation in EVERY CELL.
Facultative Heterochromatin: IS EPIGENETIC. Example is
Barr BODIES IN X chromosomes
Epigenetics include post translational modifications of Histone
Tails and Nucleotide modifications .
DNA Methylation is associated with silenced genes.
Histone Code is established by post transciptional
modifications

40. Explain the molecular steps involved in nuclear protein
import
Proteins and RNA are transported in and out of nucleus.
Nuclear Pores contains the Nuclear pore complex that appears
to fill the pore like a Stopper
Nuclear pore Complex is composed of 30 proteins called
Nucleoporins.
Nuclear Localization Signal targets a protein to the Nucleus.
For example localization of T antigen containing its Normal
Nuclear Import Signal.
40.)Explain the role of RanGTP, NLS, nucleoporin, and importin
beta in nuclear import.
Ran State can be used as a marker for Nucleus versus

Cytoplasm. Provide directionality to the system.

Ran is a small GTPase its effector molecules are located on
opposite sides of Nuclear Membrane.
Ran-GEF gives Ran GDP a GTP in the Nucleus and turns it into
Ran GTP.
RAN GAP IN Cytosol Hydrolyzes GTPase activity it hydrolyzes
third phosphate from GTP and turns it into GDP.
Ran GEF: In the nucleus, exchange nucleotide takes away a
GDP and gives it a GTP.
The distribution of these two enzymes allow protein to bind
cargo on one side and let go of it on the other side.
Nuclear Pore Complex is only looking for Cargo with Importins.
Importin B is a protein that can recognize Cargo, recognizes
Cargo because Cargo has a Nuclear Localization Signal.
Importin B recognizes NLS signal on Cargo they go to the
Nuclear pore COMPLEX and because Importins can be
recognized by Nuclear Pore Complex(Nucleoporins) they get
in. Once in the nucleus Cargo is saying to Importin let go of me
IM HERE. Importin lets go of Cargo in the Nucleus. In the
Nucleus, the Ran protein is in the GTP bond state, Ran GTP
BINDS to IMPORTIN AND IT DISPLACES CARGO. CARGO CAN
BIND IMPORTIN IN the cytoplasm because there is no RAN
GTP IN THE CYTOPLASM(ICF).
But inside the nucleus the Ran GTP is waiting for the Importin
to come in so it can bind to Importin, when it binds Importin,
Cargo falls off.
So Ran-GTP bound to Importin B and they go back out, when
they go back out there is an enzyme that hydrolyzes Ran-GTP

back to Ran GDP and lets go of Importin.

The Ran-GTP :importin B complex returns to the cytoplasm.
There the Ran GAP hydrolyzes Ran GTP TO Ran GDP which
seperates from Importin B and returns to the Nucleus



42.What are lamins, and what are their roles? How do they
contribute to human genetic disorders?
Lamins are helical proteins with globular C-terminal domains.
They are the products of multiple genes alternative Rna
processing, which is regulated in a cell development specific
fashion. Inner Surface of the Nuclear Envelope is lined by the
Nuclear Lamina.
Nuclear Lamina helps support Chromatin supports the Nuclear
Envelope and is composed of Lamins.
Lamins are also though to be associated chromatin and
involved in chromatin organization during Interphase and
Mitosis.
Defeats in Lamin protein may just affect overall structure of
the nucleus. Defeats in Lamin contributes to forms of Muscular
Dystrophy and Neurological disorders.
An example of disease related to Lamin is Progeria which is a
syndrome where children are born normal and begin to age at
an accelerated rate they die of Heart Attacks and Strokes.
Laminopathies: Mutations in Lamins that can alter the
structure of the Nucleus, the interaction between the

chromatin and the nuclear envelope, and the regulation of gene

expression(heterochromatin vs euchromatin) position of
Chromatin within the nucleus.