Catabolism of proteins Seminar No. 5 Amino acid pool ~ 80 % in muscles ~ 10 % in liver ~ 5 % in kidney ~ 5 % in blood Overview of AA metabolism Three sources of AA pool: • Proteolysis of food proteins • Proteolysis of tissue proteins • Synthesis of non-essential AA Three uses of AA pool: • Synthesis of tissue and plasma proteins • Synthesis of specialized nitrogen products • Deamination + utilisation of carbon skeleton Q. 1 A. 1 • Stomach – pepsin • Small intestine: trypsin, chymotrypsin, elastase, carboxypeptidase A/B, aminopeptidase Q. 2 A. 2 Q. 3 A. 3 L-amino acids: about seven specific transporters, symport with Na^+ D-amino acids (trace amounts): nonspecific diffusion, hydrophilic pores in membranes, D-AA cannot be utilized in the body, they are only catabolized to gain energy Q. 4 A. 4 Intracellular proteases degrade endogenous proteins, two systems: • Lysosome (non-specific degradation, no ATP) Extracellular + membrane proteins • Ubiquitin-proteasome (ATP needed) damaged/misfolded proteins, regulations proteins (with short half-life) Q. 5 A. 5 Biological value of some proteins Whey Q. 6 + 7 A. 6 + 7 • Valine (branched) • Leucine (branched) • Isoleucine (branched) • Threonine (2 C*) • Phenylalanine (aromatic ring) • Tryptophan (aromatic ring) • Lysine (basic, two NH[2] groups) • Methionine (S-CH[3]) Q. 8 A. 8 Most plant food • cereals, rice, corn (maize) – lack of Lys, Trp, Thr, Met • legumes – lack of Met Some animal food • gelatin (lack of Trp) • game, octopus, lobster (low digestibility) Conversions of AA after meal • AA from food are absorbed from intestine • Glutamate +glutamine are utilized as metabolic fuel for enterocyte • 20 % of AA in portal blood are branched AA • In liver, most AA are utilized for synthesis of proteins, Glc, FA. • Val, Leu, Ile are not metabolized in liver due to the lack of aminotrasferases TH they predominate (70 %) in central circulation • High content of ammonia in portal blood is removed by liver  urea Q. 10 A. 10 Carbon skeleton of AA is used to make FA and TAG Highly protein diet invariably leads to obesity Q. 11 A. 11 Because of lack of specific aminotransferases in liver Q. 12 A. 12 • glutamine is deaminated to glutamate [• ] glutamate + NADPH+H^+ ® glutamate semialdehyde + ADP + P[i ]• glutamate semialdehyde is transaminated to ornithine • ornithine + carbamoyl phosphate ® citrulline • citrulline is transported to kidneys where it is converted to arginine • arginine is utilized in liver for urea Citrulline is made by the addition of carbamoyl group to ornithine Q. 13 A. 13 • Deaminations of glutamine + glutamate in enterocyte • Bacterial putrefaction of proteins in the large intestine produces nitrogen catabolites (e.g. biogenic amines + ammonia), ammonia diffuses freely into portal blood TH portal blood has high concentration of NH[4]^+ TH eliminated by liver How can you decrease the production of ammonia in the human body? • Low-protein diet (especially important in liver diseases) • Alteration of colon microflora by the ingestion of: • Probiotics – live bacteria stimulating saccharolytic (fermentative) processes in large intestine instead of putrefactive ones (Lactobacillus, Bifidobacterium) – yoghurt, kefir milk • Prebiotics – non-digestible food ingredients that stimulate the growth probiotics in the colon (dietary fibre, lactulose, oligofructose, inulin) – e.g. soybeans, Jerusalem artichokes (inulin), chicory root (inulin), oats ... Ammonium ions in body fluids Conversions of AA in fasting • There is no special protein store in the body • Liver proteosynthesis is limited, proteolysis in muscles increases (insulin ↓, cortisol ↑) • The main AA released from muscles are Ala + Gln • Ala is the substrate of liver gluconeogenesis • Gln is deaminated in liver to give NH[4]^+ - urea synthesis (periportal region) • Gln is made in perivenous region – the detoxication of remaining ammonia Q. 19 ! A. 19 - Gln in muscle • Gln is released by proteolysis • Gln is product of ammonia detoxication • Gln can be viewed as a carrier of –NH[2] group from muscles to liver (periportal hepatocytes) where NH[3] is liberated and converted to urea A. 19 – Gln in enterocyte • exogenous and endogenous Gln is the source of energy for intestinal mucosa: Gln  2-OG  energy (CAC) • enterocytes have high turnover – Gln (and other AA) are needed for proteosynthesis and nucleic acid bases • limited usage of glucose and FA as fuel in enterocyte A. 19 – Gln in brain • Glutamine formation is the principal way of ammonia detoxication in CNS • Glutamine synthase reaction occurs mainly in astroglial cells • In other CNS cells is Gln the source of glumate – as the substrate for GABA Glutamine synthesis requires one mol of ATP A. 19 – Gln in liver • in periportal hepatocytes, Gln is the source of ammonia for urea synthesis • in perivenous hepatocytes, Gln is made from glutamate (Glu + NH[3] ® Gln) as the additional way of ammonia detoxication • Gln is released from liver to blood - transported to enterocytes and kidney A. 19 – Gln in kidneys • Gln is the source of energy for the kidneys, to a great extent especially in fasting and under metabolic acidosis • Gln and Glu release ammonium ions which contribute to acidic pH of urine The origin of ammonium in urine Glutaminase catalyses the hydrolysis of amide group in glutamine Multiple functions of glutamine • Synthesis of proteins • Metabolic fuel – enterocytes, lymphocytes, macrophages, fibroblasts, kidneys • Source of nitrogen in synthesis – purine, pyrimidines, NAD^+, aminosugars • Source of glutamate – GSH, GABA, ornithin, prolin, • Source of ammonium ions in urine Q. 20 A. 20 - AA in blood Resorption phase • predominate Val, Leu, Ile • liver does not take them up from circulation (no specific aminotransferases in liver for Val, Leu, Ile) Postresorption phase and fasting • predominate Gln and Ala • released from muscles (Gln + Ala) and liver (Gln) Q. 21 A. 21 - Dehydrogenation deamination of glutamate Q. 22 A. 22 Glucose-alanine cycle Q. 23 A. 23 Three ways of ammonia detoxication Q. 26 A. 26 Catabolic pathway of nitrogen (in blue colour) • dietary proteins ® AA (GIT) • transamination of AA in cells ® glutamate [• ] dehydrogenation deamination of glutamate ® NH[3 ]• detoxication of ammonia ® urea A. 26 General scheme of transamination 1. Phase of transamination 2. Phase of transamination In transaminations, nitrogen of most AA is concentrated in glutamate GMD reaction is reversible A. 27 A. 27 • AST reaction is reversible • provides aspartate for the urea synthesis Q. 28 A. 28 ammonia glutamine Q. 29 A. 29 Hb-Val-NH[2] + NH[2]-CO-NH[2] ® Hb-Val-NH-CO-NH[2] + NH[3 ] Factors affecting nitrogen balance Factors affecting nitrogen balance Q. 30 A. 30 The loss of N = 4 g/day Average content of N in proteins is 16 %. Average content of proteins in muscles is 20 %. ---------------------------------------------------------------------- 100 g prot. ........................ 16 g N x g prot. ...........................4 g N x = 400 / 16 = 25 g of proteins 100 g muscles ........................ 20 g proteins x g muscles ......................... 25 g proteins x = 2500 / 20 = 125 g of muscles