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Hanley N. Abramson
Professor of Pharmaceutical Sciences
Wayne State University
December 2009
Dynamics of Protein
And Amino Acid Metabolism
Trypsin cleaves:
Chymotrypsinogen to chymotrypsin
Proelastase to elastase
Procarboxypeptidase to carboxypeptidase
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)
Glutamate
2) NH4+ + dehydrogenase
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
9
Metabolic Classification of
the Amino Acids
• Essential and Non-essential
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
11
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
12
Glucogenic Amino Acids
• Metabolized to -ketoglutarate, pyruvate,
oxaloacetate, fumarate, or succinyl CoA
Phosphoenolpyruvate Glucose
13
Ketogenic Amino Acids
• Metabolized to acetyl CoA or acetoacetyl
CoA
5 Steps Proline
Glutamate
Glutamine
Ornithine
synthase
Urea Cycle
Arginine
Glutamine 15
Guanidino group
GABA Formation
Glutamate
decarboxylase
Glutamate Gamma-aminobutyrate
CO2 (GABA)
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
19
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
20
Synthesis of Nitric Oxide
Arginine
+ NO
Citrulline
21
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
22
Formation of Serine
Dehydrogenase
Glycolysis
Glucose
NAD+ NADH +
H+
Phosphatase
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
24
Sarcosine (N-Methylglycine)
O
O Glycine N-Methyltransferase
H2
N
H3N H3C O
O
Glycine Sarcosine
25
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)
26
Homocysteine
Homocysteinuria
• Rare; deficiency of cystathionine -synthase
• Dislocated optical lenses
• Mental retardation
• Osteoporosis
• Cardiovascular disease death
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
29
Polyamines
• Spermidine and spermine found in virtually
all procaryotic and eucaryotic cells
-Difluoromethyl-
ornithine (DFMO)
(Eflornithine) - inhibits ODC;
used to treat
Pneumocystis carinii infectons
30
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
31
Creatine and Creatinine
Creatine:
• Dietary supplement
• Used to improve athletic performance
Creatinine:
• Urinary excretion generally constant;
proportional to muscle mass
32
Histidine Metabolism:
Histamine Formation
Histidine
decarboxylase
O2 Phenylalanine-4- NADP+
Monooxygenase
(Phenylalanine
H2O NADPH + H+
hydroxylase)
Dihydrobiopterin
+
Tyrosine
(Non-essential)
34
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)
36
Sapropterin (Kuvan)
• Synthetic tetrahydrobiopterin
• Indicated for reduction of phenylalanine blood
levels in patients with BH4-responsive PKU
37
Catecholamine Biosynthesis
Catechol
Tyr hydroxylase
O2
Tyrosine Dihydroxyphenylalanine
(DOPA)
Epinephrine DOPA
(Adrenaline) decarboxylase CO2
Methyl Dopamine
transferase hydroxylase
S-Adenosyl-
homocysteine
SAM Dopamine
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
40
Tyramine
MAO
Tyramine
( blood pressure)
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
• Asymptomatic in childhood
44
Melanin Formation
Tyr hydroxylase
O2
Tyrosine DOPA
Tyrosinase
Trp
hydroxylase
Decarboxylase
O2
46
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)
47
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
48
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
50
Tryptophan Metabolism:
Biosynthesis of Nicotinic Acid
Several steps
Nicotinamide adenine
dinucleotide (NAD)
51