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Amino Acid Metabolism

Hanley N. Abramson
Professor of Pharmaceutical Sciences
Wayne State University
December 2009
Dynamics of Protein
And Amino Acid Metabolism

Dietary Proteins Digestion to Amino Acids

Transport in Blood to Cells

Protein Synthesis Functional Proteins

Amino Acids Protein Degradation In


Proteasomes Following
Tagging With Ubiquitin
Metabolites 2
Digestion of Proteins
Stomach: Pepsinogen Pepsin (max. act. pH 2)
Enteropeptidase
Small Intestine: Trypsinogen Trypsin

Trypsin cleaves:
Chymotrypsinogen to chymotrypsin
Proelastase to elastase
Procarboxypeptidase to carboxypeptidase

Aminopeptidases (from intestinal epithelia)

3
Intestinal Absorption
Amino Acids Oligopeptides
Lumen

Transport
Protein
Oligopeptides

Peptidases

Amino Acids

Blood
4
+
Incorporation of NH4 Into
Organic Compounds
+ - Carbamoyl
1) NH4 + HCO3 + 2 ATP Phosphate NH2CO2PO3-2 + 2 ADP +
Synthase I +
Carbamoyl Phosphate Pi + 2 H
(CPS-I)

TCA Cycle mitochondria

Glutamate
2) NH4+ + dehydrogenase

-Ketoglutarate NADPH + NADP+ Glutamate


H+

5
+
Incorporation of NH4 Into
Organic Compounds (Cont.)

3)
+ NH4+ + 2 ATP
Glutamate Glutamine
Synthase
Mg++

Glutamine
N of glutamine donated to other compounds
in synthesis of purines, pyrimidines,
6
and other amino acids
Biosynthesis of Amino Acids:
Transaminations
Amino Acid1 +-Keto Acid2 Amino Acid2 +-Keto Acid1

+
Glutamate
Pyridoxal phosphate (PLP)-
Dependent Aminotransferase

+
-Ketoglutarate
7
Transaminations: Role of PLP

Tautomerization

8
Transaminations
Glutamate-Pyruvate
Aminotransferase
Glutamate (Alanine Transferase ALT) -Ketoglutarate
+ +
Pyruvate Alanine
Glutamate-Oxaloacetate
Aminotransferase
(Aspartate Transferase AST)
Glutamate -Ketoglutarate
+ +
Oxaloacetate Aspartate

Blood levels of these aminotransferases, also called transaminases,


are important indicators of liver disease

9
Metabolic Classification of
the Amino Acids
• Essential and Non-essential

• Glucogenic and Ketogenic

10
Non-Essential Amino Acids
in Humans
• Not required in diet
• Can be formed from -keto acids by
transamination and subsequent reactions
• Alanine • Glycine
• Asparagine • Proline
• Aspartate • Serine
• Glutamate • Cysteine (from Met*)
• Glutamine • Tyrosine (from Phe*)
* Essential amino acids
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Essential Amino Acids in
Humans
• Required in diet
• Humans incapable of forming requisite
carbon skeleton
• Arginine* • Lysine
• Histidine* • Methionine
• Isoleucine • Threonine
• Leucine • Phenylalanine
• Valine • Tryptophan
* Essential in children, not in adults
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Glucogenic Amino Acids
• Metabolized to -ketoglutarate, pyruvate,
oxaloacetate, fumarate, or succinyl CoA
Phosphoenolpyruvate Glucose

• Aspartate • Methionine • Alanine


• Asparagine • Valine • Serine
• Arginine • Glutamine • Cysteine
• Phenylalanine • Glutamate • Glycine
• Tyrosine • Proline • Threonine
• Isoleucine • Histidine • Tryptophan

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Ketogenic Amino Acids
• Metabolized to acetyl CoA or acetoacetyl
CoA

Animals cannot convert acetyl CoA or


acetoacetyl CoA to pyruvate
• Isoleucine • Tryptophan
• Leucine * • Phenylalanine
• Lysine * • Tyrosine
• Threonine
* Leucine and lysine are only ketogenic
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Amino Acids Formed From
-Ketoglutarate
-Keto- 4 Steps
Transamination or
glutarate Glutamate
dehydrogenase

5 Steps Proline
Glutamate
Glutamine
Ornithine
synthase
Urea Cycle

Arginine
Glutamine 15
Guanidino group
GABA Formation
Glutamate
decarboxylase

Glutamate Gamma-aminobutyrate
CO2 (GABA)

GABA is an important inhibitory neurotransmitter


in the brain
Drugs (e.g., benzodiazepines) that enhance the effects
of GABA are useful in treating epilepsy
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Arginine Synthesis: The Urea Cycle

N-Acetylglutamate
synthase
CoASAc
Glutamate N-Acetylglutamate
Activates
4 Steps

Carbamoyl
CPS-I phosphate
+ -
NH4 + HCO3 NH2CO2PO3-2
Ornithine
Ornithine
Transcarbamoylase (OTC)
(mitochondria)
Citrulline
Ureido group 17
The Urea Cycle (Contd.)
Asp

Citrulline
Arginosuccinate
synthase
Arginosuccinate
Ornithine
Transcarbamoylase
(mitochondria)
Fumarate
Urea
H2NCONH2 Argino-
succinase
Ornithine TCA Cycle

Arginase
These reactions occur
in the cytosol 18
Arginine
Urea Formation
• Occurs primarily in liver; excreted by kidney
• Principal method for removing ammonia
• Hyperammonemia:
• Defects in urea cycle enzymes (CPS, OTC, etc.)
• Severe neurological defects in neonates
• Treatment:
» Stop protein intake
» Dialysis
» Increase ammonia excretion: Na benzoate, Na
phenylbutyrate, L-arginine, L-citrulline

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Blood Urea Nitrogen
• Normal range: 7-18 mg./dL
• Elevated in amino acid catabolism
• Glutamate N-acetylglutamate
CPS-1 activation
• Elevated in renal insufficiency
• Decreased in hepatic failure

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Synthesis of Nitric Oxide
Arginine

Nitric oxide synthase (NOS)

+ NO

Citrulline

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Nitric Oxide
• Cell messenger
• Implicated in a wide range of physiological
and pathophysiological events:
• Vasodilation:
• Activates guanylyl cyclase cGMP
• Nitroglycerin Glycerin + NO
• Sildenafil (Viagra): in vascular smooth muscle:
Blocks Phospho-
diesterase-5
NO cGMP GMP

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Formation of Serine
Dehydrogenase
Glycolysis
Glucose
NAD+ NADH +
H+

3 Steps 3-Phospho- 3-Phospho-


glycerate hydroxypyruvate
Pyruvate Inhibits Glutamate
Transaminase
-Ketoglutarate

Phosphatase

Serine (Ser) 3-Phosphoserine


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Conversion of Serine to Glycine
Dihydrofolate
reductase
Folate

Serine
Tetrahydrofolate
(FH4)
Serine hydroxymethyl
transferase (PLP-dep.)

Key intermediate
in biosynthesis of Glycine
purines and
formation of
thymine Important in
N5, N10-Methylene FH4
biosynthesis of heme,
porphyrins, and purines
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Sarcosine (N-Methylglycine)
O
O Glycine N-Methyltransferase
H2
N
H3N H3C O
O
Glycine Sarcosine

Sarcosine as a possible biomarker for prostate cancer:


See Nature Feb. 12, 2009

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Sulfur-Containing Amino Acids
Methionine
Synthase
(Vit. B12-dep.) + 5-Methyl
+ FH4 FH4
Methionine L-Homocysteine
(Essential)
Cystathionine
Serine
-synthase
(PLP-dep.)

Cystathionine
lyase

+
Cystathionine
-Hydroxy- Cysteine
butyrate (Non-essential)
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Homocysteine
Homocysteinuria
• Rare; deficiency of cystathionine -synthase
• Dislocated optical lenses
• Mental retardation
• Osteoporosis
• Cardiovascular disease death

High blood levels of homocysteine associated with


cardiovascular disease
• May be related to dietary folate deficiency
• Folate enhances conversion of
homocysteine to methionine
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Methionine Metabolism:
Methyl Donation
S-Adenosyl methionine
synthase

Methionine ATP

SAM
Decarboxylase S-Adenosyl
Methionine
(SAM)

Decarboxylated CO2
SAM R-H
Methyl-
transferases
+
R-CH3
S-Adenosyl
homocysteine 28
Polyamine Biosynthesis
Ornithine
decarboxylase
(ODC)
(PLP-dep.)

Ornithine Putrescine
(from urea cycle) CO2 Decarboxylated
SAM
Spermidine
synthase

Spermine 5’-Methylthio-
synthase adenosine
Spermine
5’-Methylthio-
adenosine

Decarboxylated Spermidine
SAM
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Polyamines
• Spermidine and spermine found in virtually
all procaryotic and eucaryotic cells

• Precise role undefined


• Bind to nucleic acids

• Inhibition of biosynthetic pathway:

-Difluoromethyl-
ornithine (DFMO)
(Eflornithine) - inhibits ODC;
used to treat
Pneumocystis carinii infectons
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Creatine and Creatinine
Arginine-glycine
transamidinase
(Kidney)

Glycine Ornithine
Arginine Guanidoacetate
SAM + ATP
Guanidoacetate
Methyltransferase S-Adenosyl-
(Liver) homocysteine
Creatinine + ADP
(Urine) Non-enzymatic
(Muscle)

Creatine kinase
(Muscle)
ADP
Creatine + Pi
Phosphocreatine
ATP
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Creatine and Creatinine
Creatine:
• Dietary supplement
• Used to improve athletic performance

Creatinine:
• Urinary excretion generally constant;
proportional to muscle mass

Creatinine Clearance Test:


• Compares the level of creatinine in urine (24 hrs.)
with the creatinine level in the blood
• Used to assess kidney function
• Important determinant in dosing of several drugs
in patients with impaired renal function

32
Histidine Metabolism:
Histamine Formation
Histidine
decarboxylase

Histidine CO2 Histamine


Histamine:
• Synthesized in and released by mast cells
• Mediator of allergic response: vasodilation, bronchoconstriction
(H1 receptors)
• H1 blockers: Diphenhydramine (Benadryl)
Loratidine (Claritin)
• Stimulates secretion of gastric acid (H2 receptors)
• H2 blockers: Cimetidine (Tagamet); ranitidine (Zantac)
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Phenylalanine and Tyrosine
Phenylalanine
(Essential)
Tetrahydrobiopterin
+ (BH4)

O2 Phenylalanine-4- NADP+
Monooxygenase
(Phenylalanine
H2O NADPH + H+
hydroxylase)

Dihydrobiopterin
+
Tyrosine
(Non-essential)

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Normal Utilization of Phenylalanine

Protein (~25%)
Phenylalanine
Tyrosine (~75%)

35
Phenylketonuria (PKU) Disease
• Deficiency of Phe hydroxylase
• Occurs in 1:20,000 live births in U.S.
• Seizures, mental retardation, brain
damage
• Treatment: limit phenylalanine intake
• Screening of all newborns mandated
in all states Tyr

Phe Phenylpyruvate
Transamination (urine)
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Sapropterin (Kuvan)

• Synthetic tetrahydrobiopterin
• Indicated for reduction of phenylalanine blood
levels in patients with BH4-responsive PKU

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Catecholamine Biosynthesis
Catechol
Tyr hydroxylase

O2

Tyrosine Dihydroxyphenylalanine
(DOPA)

Epinephrine DOPA
(Adrenaline) decarboxylase CO2

Methyl Dopamine
transferase hydroxylase
S-Adenosyl-
homocysteine
SAM Dopamine

DOPA, dopamine, norepinephrine, Norepinephrine 38


and epinephrine are all neurotransmitters
L-DOPA in Parkinsonism
Blood Brain

L-DOPA L-DOPA Dopamine


Blocks

Carbidopa Parkinsonism associated with


dopamine in brain through loss of
Dopamine neurons in basal ganglia.
Carbidopa + L-DOPA

Blood Brain Barrier 39


Monoamine Oxidase (MAO)
MAO
(in mitochondria)

R R’ Aldehyde
dehydrogenase
OH H Norepi
OH CH3 Epi
H H Dopamine
Urinary
MAO inhibitors (e.g., tranylcypromine) are useful
metabolite
in the treatment of depression
R=OH Vanillylmandelic acid (VMA)
Brain levels of dopamine and norepi.; also R=H Homovanillic acid (HVA)
serotonin
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Tyramine
MAO

Tyramine
( blood pressure)

• Tyramine found naturally in several types of cheese;


also beer and red wine.
• Tyramine intake can cause hypertensive crisis in
persons taking a MAO inhibitor ( norepi release)
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Catechol-O-Methyl
Transferase (COMT)

COMT

SAM S-Adenosyl-
Active homocysteine
catecholamine Inactive
metabolite
• COMT found in cytoplasm
• Terminates activity of catecholamines
• Catecholamine excretion products result from
combined actions of MAO and COMT
• Inhibitors of COMT (e.g., tolcapone) useful
in Parkinson’s disease 42
Homogentisic Acid Formation

Transamination

Tyrosine p-Hydroxyphenyl-
pyruvate
Deficient in O2
alkaptonuria p-Hydroxyphenyl-
pyruvate
Homogentisate dioxygenase
dioxygenase (ascorbate-dep.)
Cleavage of
aromatic ring CO2

O2
Fumarate + acetoacetate Homogentisate 43
Alkaptonuria
• Deficiency of homogentisate dioxygenase

• Urine turns dark on standing


• Oxidation of homogentisic acid

• Asymptomatic in childhood

• Tendency toward arthritis in adulthood

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Melanin Formation
Tyr hydroxylase

O2
Tyrosine DOPA
Tyrosinase

Melanin Highly colored


(Black polymer) polymeric
intermediates

Melanin formed in skin (melanocytes), eyes, and hair


In skin, protects against sunlight Dopaquinone
Albinism: genetic deficiency of tyrosinase
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Tryptophan Metabolism:
Serotonin Formation
Indole ring

Trp
hydroxylase
Decarboxylase
O2

Tryptophan 5-Hydroxy- CO2 5-Hydroxy-


(Trp) tryptophan tryptamine (5-HT);
Serotonin

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Serotonin
• Serotonin formed in:
• Brain (neurotransmitter; regulation of sleep, mood, appetite)
• Platelets (platelet aggregation, vasoconstriction)
• Smooth muscle (contraction)
• Gastrointestinal tract (enterochromaffin cells - major storage site)

• Drugs affecting serotonin actions used to treat:


• Depression
•Serotonin-selective reuptake inhibitors (SSRI)
• Migraine
• Schizophrenia
• Obsessive-compulsive disorders
• Chemotherapy-induced emesis

• Some hallucinogens (e.g., LSD) act as serotonin agonists

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L-Tryptophan
• Food supplement promoted for serotonin effects
• L-Tryptophan disaster (1989):
• Eosinophilia-myalgia syndrome (EMS)
• Severe muscle and joint pain
• Weakness
• Swelling of the arms and legs
• Fever
• Skin rash
• Eosinophilia
• Many hundreds of cases; several deaths
• Traced to impurities

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Serotonin Metabolism: 5-HIAA

MAO

Serotonin Dehydrogenase

Carcinoid tumors:
• Malignant GI tumor type
• Excretion of large amounts of 5-HIAA
5-Hydroxyindole acetic
acid (5-HIAA) (Urine) 49
Serotonin Metabolism:
Melatonin

2 Steps

Serotonin Melatonin
Melatonin:
• Formed principally in pineal gland
• Synthesis controlled by light, among other factors
• Induces skin lightening
• Suppresses ovarian function
• Possible use in sleep disorders
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Tryptophan Metabolism:
Biosynthesis of Nicotinic Acid

Several steps

Tryptophan Nicotinic acid (Niacin)

Nicotinamide adenine
dinucleotide (NAD)
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