Metabolism of amino acids Department of Biochemistry (J.D.) 2013 1 Intermediates of amino acid catabolism • Glucogenic (13) pyruvate and/or CAC intermediates • Ketogenic (2) = Leu, Lys acetyl-CoA + acetoacetate • Mixed (5) = Thr, Ile, Phe, Tyr, Trp 2 Intermediates of amino acid catabolism Ser,Ala, Cys,Gly, Gly,Thr, Ser, Ala, Thr, Cys, (Trp) Trp pyruvatepyruvát glucoseglukosa Ile, Leu, Lys, Thr acetyl-CoA acetoacetateacetoacetát Leu, Lys, Phe, Trp, Tyr Asp, Asn oxaloacetateoxalacetát CC Phe, Tyr fumarátfumarate 22-oxoglutarát-oxoglutarate Arg, Glu, Gln, His, Pro Asp succinylsukcinyl-CoA-CoA Ile, Val, Met, Thr 3 Alanine Transamination of alanine 2-oxoglutarát2-oxoglutarate Gluglutamate ALT H 3 C C H C OOH H 3 C C C OOH alanine aminotransferase NH 2 O alaninalanine pyruvátpyruvate 4 Glucose-alanine cycle liver muscle glucose glucose glycolysis gluconeogenesis pyruvate transport in blood pyruvate transamination transamination alanine alanine • alanine is non-toxic transport of ammonia from muscles to liver • in the liver, alanine is the substrate for gluconeogenesis 5 Alanine - summary • readily made from pyruvate (transamination) • ALT is clinically important enzyme, mainly in liver, elevated catalytic concentration in blood serum – liver diseases • Ala is released to blood mainly from muscles, together with Gln (postresorption phase) • semiessential AA (in metabolic stress) – important substrate for gluconeogenesis 6 Arginine no transamination Hydrolysis of arginine urea C H 2 C H 2 C H2 C HC OOH C H 2 C H 2 C H 2 C HC OOH NH NH NH 2 NH 2 2 ornitin C N H ornithine H 2 O N H 2 N H 2 glutamate argininearginin C O N H 2 urea 7 NO is signal molecule from arginine C H C H C H C HC OOH 2 2 2 C H 2 C H 2 C H 2 C HC OOH NH NH 2 NH NH 2 O , NA D P H C N H 2 C N O H NH 2 BH4 NH 2 NN-hydroxyarginin-hydroxyarginine Exogenous NO sources • glycerol trinitrate • isosorbide dinitrate O 2 , NA D P H • amyl nitrite • isobutyl nitrite • sodium nitroprusside C H 2 C H 2 C H 2 C HC OOH + N O NH NH 2 oxidnitric dusnatý oxide (nitroxidradical radikál) C O citrulincitrulline 8 NH 2 Synthesis of creatine (1. part) from Greek κρέας (meat) C H2 C OOH C H2 C OOH NH2 H2 N NH C glycinglycine NH guanidinoacetate C H2 C H2 C H2 C H C OOH C H2 C H2 C H2 C H C OOH H2N NH NH2 NH2 NH2 C NH arginine ornithine 9 Synthesis of creatine (2. part) N1-methylation of guanidinoacetate S-adenosylhomocysteine S-adenosylmethionine (SAM) C H2 C OOH C H2 C OOH H2 N NH H2 N N C C C H3 NH NH guanidinoacetate creatine N-methylguanidine-N-acetate 10 N2-Phosforylation of creatine C H2 C OOH O C H C OOH ATP H 2 H2 N N HO P N N C C H 3 C C H3 OH NH NH creatinekreatin creatinekreatinfosfát phosphate 11 Creatinine is a catabolite of creatine made in non-enzymatic reaction O C H C OOH non-enzymatic 2 C C H cyclization dehydratation 2 H N N 2 N N C C H3 H C C H3 NH - H O 2 NH kreatin creatine creatinine 12 Arginine - summery • semiessential AA (childhood) • the most basic AA (guanidine, pKB = 0.5) • no transamination, Arg releases ornithine + urea • Arg + Gly + Met creatine • releases NO (vasodilator) • OTC (over-the-counter) preparations in pharmacy 13 Serine Dehydratation + deamination of serine OH H C H 2 C C OOH C H 2 C C OOH H 3 C C C OOH - H 2 O N H 2 N H 2 N H enamineenamin imineimin H 2 O + H C C C OOH N H 3 3 O pyruvatepyruvát 14 Conversion of serine to glycine C H C OOH + H OC H F H C H 2 C H C OOH + F H4 2 2 4 NH OH NH 2 2 serinserine glycineglycin H2O + N5N10-CH2-FH4 cofactor: methylene FH4 tetrahydrofolate (FH4) 15 Transamination of serine and glucose formation 2-oxoglutarate2-oxoglutarát Glu C OOH C OOH C OOH NADH + H + H 2 N C H C O C H OH C H 2 OH C H 2 OH C H 2 OH serineserin hydroxypyruvatehydroxypyruvát glycerátglycerate ATP reverse reaction: synthesis of serine ADP pathway is different - through phosphoserine C OOH 3-P-glycerát3-P-glycerate glucoseglukosa C H OH O C H 2 O P O 16 O Decarboxylation of serine gives ethanolamine. Methylation of ethanolamine leads to choline decarboxylation methylation C H3 HO C H2 C H C OOH HO C H2 C H2 NH2 HO C H2 C H2 N C H3 NH2 C H3 serine ethanolamine choline Betaine is made by choline oxidation C H 3 C H 3 oxidation H O C H 2 C H 2 N C H 3 H OOC C H 2 N C H 3 C H 3 C H 3 choline betaine 17 Serine - summary • non-essential glucogenic AA • source of C1 fragments (attached to tetrahydrofolate) • component of glycerophospholipids • decarboxylation gives ethanolamine choline • carbon skeleton used for selenocysteine • serine side chain in proteins: the site of phosphorylation the linkage of oligosaccharides (O-glycoside bond) nucleophilic -OH group in active site of enzyme (serine proteases) 18 Glycine The complete catabolism of glycine + N H C H 2 C O O H + F H 4 N 5 N 1 0 C H 2 F H 4 + C O 2 3 N H 2 C1 fragment (methylene) is transferred to tetrahydrofolate 19 Oxidative deamination of glycine O O C C oxid. HO OH H O O O 2 oxalateoxalát C H 2 C OOH C H C OOH C C - NH 3 NH 2 NH H OH F A D F A D H 2 glyoxalát glyoxylate O - C O 2 C S C oA H H 2 O 2 O 2 formyl-CoA 60 % catabolism of glycine and ethanolamine 30 % catabolism of vitamin C 10 % food (spinach, rhubarb, mangold, tea, cocoa) 20 Glycine - summary Catabolism Anabolic conversions • complete oxidation to • donor of C1 fragment • serine CO2 + NH3 • porphyrines • oxidative deamination • purine bases to oxalate • creatine • glutathione (GSH) • conjugation agent (bile acids, xenobiotics) 21 Threonine no transamination Threonine (4C) is split to glycine (2C) and acetaldehyde (2C) C OOH C OOH serinserine pyruvátpyruvate H 2 N C H 2 H 2 N C H glycineglycin H C OH O O C H 3 C H C C H 3 C 3 H S C oA acetaldehydacetaldehyde acetyl-CoA • essential AA • two asymmetric C atoms • the site of phosphorylation and glycosylation in proteins 22 Methionine no transamination Methionine is methylation agent (homocysteine side product) ethanolamine PP i + P i noradrenaline C H substratesubstrát 3 guanidinacetate HOOC C H C H 2 C H 2 S Rib A d ATP NH 2 S-Sadenosylmethionine-adenosylmethionin substratesubstrát -CCHH 33 C H 3 cholinecholin HOOC C H C H 2 C H 2 S adrenalinadrenaline HOOC C H C H 2 C H 2 S Rib A d creatinekreatin NH 2 NH 2 S -adenosylhomocystein methioninemethionin remethylationremethylace HOOC C H C H C H S H F H 4 2 2 B NH 2 12 homocysteinehomocystein 23 C H 3 F H 4 S-Adenosylmethionine (SAM) contains trivalent positively charged sulfur atom NH2 N N N N C H 3 S HOOC C H C H2 C H2 O NH 2 cation - sulfonium OH OH 24 Cysteine is made from methionine methionine HOOC C H C H 2 C H 2 S H NH 2 homocysteinehomocystein pyridoxal-P kondenzacecondensation se serinem with serine H 2 O HOOC C H C H 2 C H 2 S C H 2 C H C OOH NH 2 NH 2 cystathioninecystathionin C H2 C H C OOH odštěpenícysteine cysteinu release S H NH2 HOOC C H C H 2 C H 2 OH B12 cysteine succinyl-CoA NH 2 homoserinehomoserin 25 Methionine - summery • essential AA, rather rare in foodstufs • S-adenosylmethionine (SAM) is methylation agent • metabolized to cysteine Cys is non-essential AA • C-skeleton of cysteine comes from serine, sulfur atom from methionine • final catabolite is succinyl-CoA (glucogenic) 26 Homocysteine is harmful • mechanism of its action is not yet understood • direct action on blood vessel epithelium • decreases thrombocyte life and fibrinolysis • supports formation of oxygen radicals – damage of vessel wall • increases LDL lipoperoxidation • elevated blood level of homocysteine is risk factor of cardiovascular diseases to eliminate homocysteine - three vitamins are needed: folate, cobalamine, pyridoxin 27 Cysteine Cysteine catabolism: oxygenation of -SH group C OOH C OOH C OOH H 2 N C H H N C H oxygenationoxygenace oxygenation 2 H 2 N C H C H 2 C H 2 O 2 C H 2 S H S O S O O OH OH cysteinecystein cysteinecysteinsulfinát sulfinate cysteovácysteic kyselina acid dekarboxylacedecarboxylation transaminationtransaminace C OOH H 2 N C H 2 oxygenationoxidace H 2 N C H 2 O C C H 2 C H 2 C H S O S O 2 O OH S OH O OH hypotaurinehypotaurin taurinetaurin sulfinylpyruvátsulfinylpyruvate 28 The formation of sulfite C OOH C OOH hydrolytic hydrolytické O C O C odštěpenícleavage sulfitu of sulfite C H 3 H 2 O C H 2 pyruvátpyruvate S O OH OH O sulfinylpyruvátsulfinylpyruvate + 2 H S S O OH O O sulfit (siřičitan)sulfite - under physiol. pH – dissociation only to HSO3 29 Sulfite oxidase catalyzes sulfate formation NH 2 N O N O S O O N N O P O O O cysteine PAPS O O OH P O O - 2- + - HSO3 + H2O SO4 + 3H + 2e blood plasma acidify (0.5 mmol/l) body fluids reduce molybdopterine 30 urine Distinguish 2- Sulfite anion SO3 Sulfide inorganic anion S2- (e.g.