Transcript Lecture 27
FCH 532 Lecture 22 Chapter 26: Amino acid metabolism Quiz Monday on Transamination mechanism Quiz on Wed. for Urea Cycle Page 987 Urea Cycle • • • • • • • Excess nitrogen is excreted after the metabolic breakdown of amino acids in one of three forms: Aquatic animals are ammonotelic (release NH3 directly). If water is less plentiful, NH3 is converted to less toxic products, urea and uric acid. Terrestrial vertebrates are ureotelic (excrete urea) Birds and reptiles are uricotelic (excrete uric acid) Urea is made by enzymes urea cycle in the liver. The overall reaction is: NH3+ NH3 + HCO3- + -OOC-CH2-CH-COOAsp O 3ATP 2ADP + 2Pi + AMP + PPi NH2-C-NH2 + -OOC-CH=CH-COOUrea Fumarate Urea Cycle • • • 2 urea nitrogen atoms come from ammonia and aspartate. Carbon atom comes from bicarbonate. 5 enzymatic reactions used, 2 in the mitochondria and 3 in the cytosol. NH3+ NH3 + HCO3- + -OOC-CH2-CH-COOAsp O 3ATP 2ADP + 2Pi + AMP + PPi NH2-C-NH2 + -OOC-CH=CH-COOUrea Fumarate Page 992 Carbamoyl phosphate synthetase • • Carbamoyl phosphate synthetase (CPS) catalyzes the condensation and activation NH3 and HCO3- to form carbomyl phosphate (first nitrogen containing substrate). Uses 2 ATPs. O 2ATP + NH3 + HCO3- NH2-C-OPO3- + 2ADP + 2Pi Carbamoyl phosphate • 1. 2. Eukaryotes have 2 types of CPS enzymes Mitochondrial CPSI uses NH3 as its nitrogen donor and participates in urea biosynthesis. Cytosolic CPSII uses glutamine as its nitrogen donor and is involved in pyrimidine biosynthesis. Figure 26-8 The mechanism of action of CPS I. • 1. Page 993 CPSI reaction has 3 steps Activation of HCO3- by ATP to form carboxyphosphate and ADP. 2. Nucelophilic attack of NH3 on carboxyphosphate, displacing the phsophate to form carbamate and Pi. 3. Phosphorylation of carbamate by the second ATP to form carbamoyl phosphate and ADP The reaction is irreversible. Allosterically activated by Nacetylglutamate. Figure 26-9 X-Ray structure of E. coli carbamoyl phosphate synthetase (CPS). • • • • Page 993 • E. coli has only one CPS (homology to CPS I and CPS II) Heterodimer (inactive). Allosterically activated by ornithine (heterotetramer of (4). Small subunit hydrolyzes Gln and delivers NH3 to large subunit. Channels intermediate of two reactions from one active site to the other. Page 992 Ornithine transcarbomylase • • • • Transfers the carbomoyl group of carbomyl phosphate to ornithine to make citrulline Reaction occurs in mitochondrion. Ornithine produced in the cytosol enters via a specific transport system. Citrulline is exported from the mitochondria. Page 992 Arginocuccinate Synthetase • • • • 2nd N in urea is incorporated in the 3rd reaction of the urea cycle. Condensation reaction with citrulline’s ureido group with an Asp amino group catalyzed by arginosuccinate synthetase. Ureido oxygen is activated as a leaving group through the formation of a citrulyl-AMP intermediate. This is displaced by the Asp amino group to form arginosuccinate. Page 994 Figure 26-10 The mechanism of action of argininosuccinate synthetase. Arigininosuccinase and Arginase • • • • • • Argininosuccinse catalyzes the elimination of Arg from the the Asp carbon skeleton to form fumurate. Arginine is the immediate precursor to urea. Fumurate is converted by fumarase and malate dehydrogenase to to form OAA for gluconeogenesis. Arginase catalyzes the fifth and final reaction of the urea cycle. Arginine is hydrolyzed to form urea and regenerate ornithine. Ornithine is returned to the mitochondria. Page 992 1. Carbamoyl phosphate synthetase (CPS) 2. Ornithine transcarbamoylase 3. Argininosuccinate synthetase 4. Arginosuccinase 5. Arginase Regulation of the urea cycle • • • • • Carbamoyl phosphate synthetase I is allosterically activated by N-acetylglutamate. N-acetylglutamate is synthesized from glutamate and acetylCoA by N-acetylglutamate synthase, it is hydrolyzed by a specific hydrolase. Rate of urea production is dependent on [N-acetylglutamate]. When aa breakdown rates increase, excess nitrogen must be excreted. This results in increase in Glu through transamination reactions. Excess Glu causes an increase in N-acetylglutamate which stimulates CPS I causing increases in urea cycle. Metabolic breakdown of amino acids • • • • Degradation of amino acids converts the to TCA cycle intermediates or precursors to be metabolized to CO2, H2O, or for use in gluconeogenesis. Aminoacids are glucogenic, ketogenic or both. Glucogenic amino acids-carbon skeletons are broken down to pyruvate, -ketoglutarate, succinyl-CoA, fumarate, or oxaloacetate (glucose precursors). Ketogenic amino acids, are broken down to acetyl-CoA or acetoacetate and therefore can be converted to fatty acids or ketone bodies. Metabolic breakdown of amino acids • • • • Degradation of amino acids converts the to TCA cycle intermediates or precursors to be metabolized to CO2, H2O, or for use in gluconeogenesis. Aminoacids are glucogenic, ketogenic or both. Glucogenic amino acids-carbon skeletons are broken down to pyruvate, -ketoglutarate, succinyl-CoA, fumarate, or oxaloacetate (glucose precursors). Ketogenic amino acids, are broken down to acetyl-CoA or acetoacetate and therefore can be converted to fatty acids or ketone bodies. Page 995 Figure 26-11 Degradation of amino acids to one of seven common metabolic intermediates. Regulation of the urea cycle • • • • • Carbamoyl phosphate synthetase I is allosterically activated by N-acetylglutamate. N-acetylglutamate is synthesized from glutamate and acetylCoA by N-acetylglutamate synthase, it is hydrolyzed by a specific hydrolase. Rate of urea production is dependent on [N-acetylglutamate]. When aa breakdown rates increase, excess nitrogen must be excreted. This results in increase in Glu through transamination reactions. Excess Glu causes an increase in N-acetylglutamate which stimulates CPS I causing increases in urea cycle. Metabolic breakdown of amino acids • • • • Degradation of amino acids converts the to TCA cycle intermediates or precursors to be metabolized to CO2, H2O, or for use in gluconeogenesis. Aminoacids are glucogenic, ketogenic or both. Glucogenic amino acids-carbon skeletons are broken down to pyruvate, -ketoglutarate, succinyl-CoA, fumarate, or oxaloacetate (glucose precursors). Ketogenic amino acids, are broken down to acetyl-CoA or acetoacetate and therefore can be converted to fatty acids or ketone bodies. Metabolic breakdown of amino acids • • • Glucogenic amino acids - Ala, Ser, Cys, Gly, Met, Arg, Gln, Glu, Asn, Asp, Pro, His, Val Ketogenic amino acids - Leu, Lys Glucogenic/Ketogenic amino acids - Ile, Phe, Thr, Trp, Tyr Pathways can be organized into groups degraded into the the seven metabolic intermediates: pyruvate, oxaloacetate, aketoglutarate, succinyl-CoA, fumarate, acetyl-CoA and acetoacetate. Acetoacetyl-CoA can be directly converted to acetyl-CoA. Page 995 Figure 26-11 Degradation of amino acids to one of seven common metabolic intermediates. Ala, Cys, Gly, Ser, Thr are degraded to pyruvate • • Trp can also be included since its breakdown product is Ala. Alanine is converted to pyruvate through a transamination reaction which transfers the amino group to -ketoglutarate to form glutamate and pyruvate. Alanine aminotransferase 2. Serine dehydratase 3. Glycine cleavage system 4, 5.Serine hydroxymethyltransferase 6. Threonine dehydrogenase 7. -amino-ketobutyrate lyase. Page 996 1. Alanine aminotransferase 2. Serine dehydratase 3. Glycine cleavage system 4, 5.Serine hydroxymethyltransferase 6. Threonine dehydrogenase 7. -amino-ketobutyrate lyase. Page 996 1. Serine dehydratase • • • PLP-enzyme forms a PLP-amino acid Schiff base (like transamination) catalyzes removal of the amino-acid’s hydrogen. Substrate loses the -OH group undergoing an elimination of H2O rather than deamination. Aminoacrylate, the product of this dehydration reaction, tautomerizes to the imine which hydrolyzes to pyruvate and ammonia. Page 997 Figure 26-13 The serine dehydratase reaction. 1. Formation of Ser-PLP Schiff base, 2. Removal of the -H atom of serine, 3. elimination of OH-, 4. Hydrolysis of Schiff base, 5. Nonenzymatic tautomerization to the imine, 6. Nonenzymatic hydrolysis to form pyruvate and ammonia. Alanine aminotransferase 2. Serine dehydratase 3. Glycine cleavage system 4, 5.Serine hydroxymethyltransferase 6. Threonine dehydrogenase 7. -amino-ketobutyrate lyase. 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