Lecture 11_AA Metabolism PDF

PHRM246 Amino Acid Metabolism Dr S Naidoo/ BPharm/Pharm Sc/Health Sc/UKZN 2 3 4 5 6 7  Dynamics of Protein And Amino Acid Metabolism Dietary Proteins Digestion to Amino Acids Transport in Blood to Cells Protein Synthesis Funct...

PHRM246 Amino Acid Metabolism Dr S Naidoo/ BPharm/Pharm Sc/Health Sc/UKZN 2 3 4 5 6 7  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 8  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)  Intestinal Absorption Amino Acids Oligopeptides Lumen Transport Protein Oligopeptides Peptidases Amino Acids Blood 10  + 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 O NH3+ 2) NH4+ + - O 2 CCH 2 CH 2 CCO 2 - dehydrogenase - O 2 CCH 2 CH 2 CHCO 2 - a-Ketoglutarate NADPH + NADP+ Glutamate H+ 11  + Incorporation of NH4 … 3) NH3+ + - O 2 CCH 2 CH 2 CHCO 2 - + NH4 + 2 ATP Glutamate Glutamine Synthase Mg++ O NH3 + H2 NCCH 2 CH 2 CHCO 2 - Glutamine N of glutamine donated to other compounds in synthesis of purines, pyrimidines, and other amino acids 12  Biosynthesis of Amino Acids: Transaminations Amino Acid1 +a-Keto Acid2 Amino Acid2 +a-Keto Acid1 NH3+ O - O 2 CCH 2 CH 2 CHCO 2 - + R-CCO - 2 Glutamate Pyridoxal phosphate (PLP)- Dependent Aminotransferase O NH2 - O 2 CCH 2 CH 2 CCO 2 - + R-CHCO 2 - a-Ketoglutarate 13  Transaminations Glutamate-Pyruvate Aminotransferase Glutamate (Alanine Transferase ALT) a-Ketoglutarate + + Pyruvate Alanine Glutamate-Oxaloacetate Aminotransferase (Aspartate Transferase AST) Glutamate a-Ketoglutarate + + Oxaloacetate Aspartate Blood levels of these aminotransferases, also called transaminases, are important indicators of liver disease 15 Metabolic Classification of the Amino Acids Essential and Non-essential Glucogenic and Ketogenic 16  Non-Essential Amino Acids in Humans Not required in diet Can be formed from a-keto acids by transamination and subsequent reactions Alanine Glycine Asparagine Proline Aspartate Serine Glutamate Cysteine (from Met*) Glutamine Tyrosine (from Phe*) * Essential amino acids 17  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 18  Glucogenic Amino Acids Metabolized to a-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 19  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 20  Amino Acids Formed From a-Ketoglutarate O - O 2 CCH 2 CH 2 CCO 2 - a-Keto- 4 Steps Transamination or glutarate Glutamate dehydrogenase + CO2 - N NH3+ - - 5 Steps H H Proline O 2 CCH 2 CH 2 CHCO 2 Glutamate NH3+ + Glutamine H3 NCH 2 CH 2 CH 2 CHCO 2 - Ornithine synthase Urea Cycle O NH3 + NH2 NH3+ + H2 NCCH 2 CH 2 CHCO 2 - H2 N=C-HNCH 2 CH 2 CH 2 CHCO 2 - Arginine Glutamine 21 Guanidino group  GABA Formation Glutamate NH3+ decarboxylase NH3 + - O 2 CCH 2 CH 2 CHCO 2 - - O 2 CCH 2 CH 2 CH 2 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 22 2 Ammonia + carbon dioxide + 3ATP ---> urea + water + 3 ADP 23 Arginine Synthesis: The Urea Cycle NH3+ N-Acetylglutamate NHCOCH3 - - synthase O 2 CCH 2 CH 2 CHCO 2 - O 2 CCH 2 CH 2 CHCO 2 - CoASAc Glutamate N-Acetylglutamate Activates 4 Steps Carbamoyl NH3+ CPS-I phosphate + + NH4 + HCO3 - NH2CO2PO3-2 H3 NCH 2 CH 2 CH 2 CHCO 2 - Ornithine NH3+ Ornithine Transcarbamoylase (OTC) NH2 CONHCH 2 CH 2 CH 2 CHCO 2 - (mitochondria) Ureido group Citrulline 24  The Urea Cycle… + Asp NH3+ NH3 -CHCH2CO2 - NH3+ + - NH2 CONHCH 2 CH 2 CH 2 CHCO 2 CO2 - H2 N=C-HNCH 2 CH 2 CH 2 CHCO 2 - Citrulline Arginosuccinate NH-CHCH2CO2- synthase CO2- Arginosuccinate Ornithine Transcarbamoylase (mitochondria) H CO2 - Fumarate NH3+ Urea + - O2 C H H3 NCH 2 CH 2 CH 2 CHCO 2 - H2NCONH2 Argino- succinase Ornithine TCA Cycle Arginase NH2 NH3+ + These reactions occur H2 N=C-HNCH 2 CH 2 CH 2 CHCO 2 - in the cytosol 25 Arginine  Urea Formation Occurs primarily in liver; excreted by kidney Principal method for removing ammonia Hyper-ammonemia: Defects in urea cycle enzymes Severe neurological defects in neonates Treatment: » Stop protein intake » Dialysis » Increase ammonia excretion: Na benzoate, Na phenylbutyrate, L-arginine, L-citrulline 26  Blood Urea Nitrogen (BUN) 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 Elevated BUN (uremia) in both acute and chronic renal (kidney) failure. Various diseases damage the kidney and cause faulty urine formation and excretion. Congestive heart failure leads to low bp & reduced renal filtration, therefore, BUN Urinary tract obstructions can also lead to an increased BUN. Severe case -hemo-dialysis removes the soluble urea and other waste products from the blood. Ions, such as Na+ and Cl- are maintained at the same concentration in the dialyzing solution, therefore - no net diffusion occurs out of circulation. Synthesis of Nitric Oxide NH2 NH3+ + Arginine H2 N=C-HNCH 2 CH 2 CH 2 CHCO 2 - Nitric oxide synthase (NOS) NH3+ NH2 CONHCH 2 CH 2 CH 2 CHCO 2 - + NO (vasodilator) Citrulline 28  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 29  Formation of Serine CO 2 - CO 2 - Dehydrogenase Glycolysis Glucose H C OH C=O NAD+ NADH + H+ CH2 OPO3-2 CH2 OPO3-2 3 Steps 3-Phospho- 3-Phospho- glycerate hydroxypyruvate Pyruvate Inhibits Glutamate Transaminase a-Ketoglutarate - CO 2 CO 2 - Phosphatase H C NH3 + H C NH3 + CH2 OH CH2 OPO3-2 Serine (Ser) 3-Phosphoserine 30 Conversion of Serine to Glycine H Dihydrofolate H2 N N N reductase CO 2 - Folate N N CH 2 NHR OH H C NH3 + Serine H Tetrahydrofolate CH2 OH (FH4) Serine hydroxymethyl transferase (PLP-dep.) H N Key intermediate CO 2 - in biosynthesis of N CH 2 Glycine purines and H C NH3 + N formation of H2 C H thymine Important in N5, N10-Methylene FH4 biosynthesis of heme, porphyrins, and purines 31  Sarcosine (N-Methylglycine) O O Glycine N-Methyltransferase H2 N H3N H3C O O Glycine Sarcosine Sarcosine as a possible biomarker for prostate cancer (Nature Feb. 12, 2009) 32  Sulfur-Containing Amino Acids Methionine Synthase NH3+ NH3 + (Vit. B12-dep.) HSCH 2CH 2CHCO 2 + 5-Methyl - - + FH4 CH 3SCH 2CH 2CHCO 2 FH4 Methionine L-Homocysteine (Essential) CO 2 - Cystathionine Serine b-synthase H C NH3 + (PLP-dep.) CH2 OH Cystathionine NH3+ OH NH3+ lyase - SCH 2CH 2CHCO 2 CH 3CHCH 2CO 2 - + HSCH 2CHCO 2- CH2CHCO2- Cystathionine b-Hydroxy- Cysteine NH3 + butyrate (Non-essential) 33  Homocysteine Homocysteinuria Rare; deficiency of cystathionine b-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 34  Methionine Metabolism: Methyl Donation NH2 N N S-Adenosyl methionine N N NH3 + synthase + - - O2 CCHCH2 CH2 -S-H2 C O CH 3SCH 2CH 2CHCO 2 ATP NH3 + CH3 Methionine OH OH SAM Decarboxylase S-Adenosyl Methionine NH2 (SAM) NH2 N N N Decarboxylated N CO2 SAM R-H N N N Methyl- N - transferases + + O2 CCHCH2 CH2 -S-H2 C O H3 NCH2 CH2CH 2-S-H2C O NH3 + R-CH3 CH3 S-Adenosyl OH OH 35 homocysteine OH OH  Polyamine Biosynthesis Ornithine decarboxylase NH3+ (ODC) + + (PLP-dep.) + NH3 H3 NCH 2 CH 2 CH 2 CHCO 2 - H3N Ornithine Putrescine (from urea cycle) CO2 Decarboxylated SAM Spermidine H synthase + + + N+ NH3 H3N N 5’-Methylthio- H Spermine H H adenosine synthase Spermine 5’-Methylthio- H + + +N NH3 adenosine H3N H Decarboxylated Spermidine SAM 36  Polyamines Spermidine and spermine found in virtually all prokaryotic and eukaryotic cells Precise role undefined Bind to nucleic acids Inhibition of biosynthetic pathway: CO2H a-Difluoromethyl- NH2 ornithine (DFMO) H2N (Eflornithine) - inhibits ODC; CHF2 used to treat Pneumocystis carinii infectIons 37  Creatine and Creatinine Arginine-glycine transamidinase NH2 NH2 NH3+ (Kidney) + + H2 N=C-HNCH 2 CH 2 CH 2 CHCO 2 - H2 N=C-HNCH 2 CO 2 - Glycine Ornithine Arginine Guanidoacetate SAM + ATP H Guanidoacetate N O Methyltransferase S-Adenosyl- (Liver) homocysteine Creatinine HN + ADP (Urine) N Non-enzymatic (Muscle) NHPO3-2 CH3 + Creatine kinase H2 N=C-NCH 2 CO 2 - NH2 (Muscle) + CH3 H2 N=C-NCH 2 CO 2 - ADP Creatine Phosphocreatine + Pi CH3 ATP 38  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 39  Histidine Metabolism: Histamine Formation + NH3 H N CH2CHCO2- Histidine H decarboxylase N CH2CH2NH2 N Histidine CO2 N 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) 40  Phenylalanine and Tyrosine Phenylalanine H H2 N N N H (Essential) Tetrahydrobiopterin H NH3+ + HN (BH4) N CHCHCH 3 CH2CHCO2- O H HO OH O2 Phenylalanine-4- NADP+ Monooxygenase (Phenylalanine H2O NADPH + H+ hydroxylase) HO N N H2N NH3+ Dihydrobiopterin + CH2CHCO2- HN N CHCHCH 3 Tyrosine O H HO OH (Non-essential) 41 Normal Utilization of Phenylalanine Protein (~25%) Phenylalanine Tyrosine (~75%) 42 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) O 43 CH2CCO2-  Catecholamine Biosynthesis HO Catechol HO Tyr hydroxylase NH3 + NH3+ CH2CHCO2- O2 HO CH2CHCO2- Tyrosine Dihydroxyphenylalanine (DOPA) HO DOPA Epinephrine (Adrenaline) decarboxylase CO2 HO CHCH2NHCH3 HO OH Methyl Dopamine transferase hydroxylase S-Adenosyl- HO HO CH2 CH2 NH2 homocysteine SAM Dopamine HO CHCH2NH2 DOPA, dopamine, norepinephrine, Norepinephrine 45 OH and epinephrine are all neurotransmitters  L-DOPA in Parkinsonism Blood Brain L-DOPA L-DOPA Dopamine HO CH3 Blocks HO CH2 -C-CO2 H Carbidopa NHNH2 Parkinsonism associated with dopamine in brain through loss of Dopamine neurons in basal ganglia. Carbidopa + L-DOPA Blood Brain Barrier 46  Monoamine Oxidase (MAO) HO MAO HO (in mitochondria) HO CHCH2 NHR' CHCHO HO R R R R’ Aldehyde OH H Norepinephrine dehydrogenase OH CH3 Epinephrine HO H H Dopamine HO CHCO2 H Urinary R MAO inhibitors (e.g., tranylcypromine) are useful metabolite in the treatment of depression Brain levels of dopamine and norepineph.; also R=OH Vanillylmandelic acid (VMA) serotonin R=H Homovanillic acid (HVA)  Tyramine OH OH MAO CH2 CH2 NH2 CH2 CHO 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 ( nor-epi release) 48  Catechol-O-Methyl Transferase (COMT) HO HO COMT HO CHCH2 NHR' CH3 O CHCH2 NHR' SAM S-Adenosyl- Active R homocysteine R 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 49 Homogentisic Acid Formation HO HO Transamination NH3 + O CH2CHCO2- CH2 CCO2- Tyrosine p-Hydroxyphenyl- pyruvate Deficient in O2 alkaptonuria p-Hydroxyphenyl- pyruvate OH Homogentisate dioxygenase dioxygenase (ascorbate-dep.) Cleavage of aromatic ring CO2 O2 CH2 CO2- Fumarate + acetoacetate OH Homogentisate 50  Alkaptonuria Deficiency of homogentisate dioxygenase Urine turns dark on standing Oxidation of homogentisic acid Asymptomatic in childhood Tendency toward arthritis in adulthood 51  Melanin Formation HO HO Tyr hydroxylase NH3+ NH3+ CH2CHCO2- O2 HO CH2CHCO2- Tyrosine DOPA Tyrosinase O Melanin Highly colored (Black polymer) polymeric intermediates O CH2 CHCO2 - NH3 Melanin formed in skin (melanocytes), eyes, and hair + In skin, protects against sunlight Dopaquinone Albinism: genetic deficiency of tyrosinase 52  Tryptophan Metabolism: Serotonin Formation + Indole ring NH3 + NH3 CH2 CHCO2 - CH2 CHCO2 - CH2CH2NH2 Trp hydroxylase HO HO Decarboxylase N O2 H N N H H Tryptophan 5-Hydroxy- CO2 5-Hydroxy- (Trp) tryptophan tryptamine (5-HT); Serotonin 53  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 54  L-Tryptophan Food supplement promoted for serotonin effects 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 55  Serotonin Metabolism: 5-HIAA CH2CH2NH2 CH2CHO HO MAO HO N N H H Serotonin Dehydrogenase CH2 CO2H HO Carcinoid tumors: Malignant GI tumor type N H Excretion of large amounts of 5-HIAA 5-Hydroxyindole acetic acid (5-HIAA) (Urine) 56  Serotonin Metabolism: Melatonin CH2CH2NH2 CH2CH2NHCOCH3 HO H3CO 2 Steps N N H H 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 57  Tryptophan Metabolism: Biosynthesis of Nicotinic Acid + NH3 CH2 CHCO2 - CO2H Several steps N N H Tryptophan Nicotinic acid (Niacin) Nicotinamide adenine dinucleotide (NAD) 58