Proteins and Amino acids metabolism, Slides of Medical Biochemistry

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Protein and Amino Acid Metabolism Medical Biochemistry SoM&D – UDOM AMINO ACID OXIDATION AND UREA BIOSYNTHESIS Dietary Protein Degradation • Pepsin digests ingested proteins to smaller peptides – Acts on N-terminal side of Leu, Phe, Trp, Tyr • Passage of the acid stomach contents triggers secretin hormone secretion into the blood • Secretin stimulates secretion of HCO3 - into the small intestine by the pancreas neutralizing gastric acid Dietary Protein Degradation • Arrival of AA in the duodenum triggers release of the cholecystokinin hormone into the blood • Cholecystokinin stimulates secretion of zymogens trypsinogen, chymotrypsinogen, procarboxypeptidases A and B by the pancreatic exocrine cells • Enteropeptidase, a proteolytic enzyme secreted by intestinal cells converts trypsinogen to trypsin Dietary Protein Degradation • Trypsin converts other trypsinogens and other zymogens to their active forms • Trypsin and chymotrypsin further degrades the peptides, at sites different from those of pepsin, into even shorter peptides Cellular Protein Degradation • There are two major cellular mechanisms for degrading damaged or unneeded proteins in the cell. – ATP dependent ubiquitin-proteosome mechanism. This mainly degrades proteins that were synthesized within the cell (endogenous) – The non-energy-dependent lysosomal degradation. Lysosomes primarily degrades extracellular proteins, eg plasma proteins, that enter the cell by endocytosis. • Membrane and some long lived cellular proteins Cellular Protein Degradation • Proteins destined for degradation by the ubiquitin-proteosome mechanism are first covalently attached to ubiquitin, a small globular protein • Ubiquitin functions as a tag on these proteins enabling proteosome, a barrel shaped protein with proteolytic activity, to recognize and act on them. – A 76 amino acid protein – Covalently attached to proteins via lysine residues – Protein degradation, trafficking Cellular Protein Degradation • Proteosome cuts the target protein into fragments that are then further degraded to AA. The process is energy dependent • Protein degradation is not random. It is influenced by some structural aspects of the protein. – Eg. Alteration by oxidation, ubiquitination Fate of Amino Acids – Excess/unused AA are degraded • Deamination of AA to form α-keto acids, the carbon skeletons of amino acids • The Amino groups (nitrogen) can by donated/used in biosynthetic processes such as purine and pyrimidine biosynthesis or disposed as urea in ureotelic organisms (animals), as uric acid in uricotelic organisms (reptiles and birds) or ammonia in ammonotelic organisms (fishes) • The carbon skeleton join the TCA cycle and oxidized to CO2 and H2O or provide 3C and 4C intermediates for gluconeogenesis Fate of Amino Acids • In animals AA are oxidatively degraded in the following metabolic circumstances: – During the normal synthesis and degradation of cellular proteins (protein turnover) the amino acids that are released and are not needed for new protein synthesis – When the ingested amino acids exceed the body’s needs for synthesis of proteins and other AA products, the surplus is catabolized (amino acids cannot be stored) – During starvation or in uncontrolled diabetes mellitus cellular proteins are used as fuel. Interorgan Amino Acid Exchange • Interorgan amino acid transport is a highly active and regulated process • It provides amino acids to all tissues of the body, both for protein synthesis and to enable amino acids to be used for specific metabolic functions • It is also an important component of plasma amino acid homeostasis Interorgan Amino Acid Exchange Kidney Urea Glucose Interorgan Amino Acid Exchange Brain Glutamine Valine, Isoleucine Alanine Alanine & Go Urea } Cells of the —> Lactate Glucose immune system TAZ Amino acids a-Keto acids Glutamine Amino Acid Catabolism • Unlike the catabolism of glucose or fatty acids, all amino acids have an amino group • AA degradation involves the key step of separating the amino groups from the carbon skeletons – deamination • The amino groups are channeled into amino group metabolism pathways – Used in biosynthetic processes – Nitrogen excretion Amino Acid Catabolism • Amino acids derived from dietary proteins are the source of most amino groups • Most amino acids are metabolized in the liver – Excess amino groups (excess nitrogen) are channeled into the urea cycle in hepatocytes and subsequently excreted as urea by kidneys • Excess amino groups/ammonia generated in extrahepatic tissues are collected and transported from those tissues to the liver and kidney as amino groups of glutamate, glutamine and alanine Amino Acid Catabolism • Four amino acids: Glu, Gln, Ala, Asp play central roles in nitrogen metabolism • These 4 AA are also most easily converted to TCA cycle intermediates • Glu and Gln are especially important, they serve as “collectors” of amino groups from other amino acids – In the cytosol of hepatocytes, amino groups from most amino acids are transferred to α-ketoglutarate to form glutamate – Some amino acids are metabolised to glutamate Amino Acid Catabolism Amino acids from FIGURE 18-2 Amino group c ingested protein of amino groups (shaded) in ve trogen. Excess NH is excrete Cellular Liver urea (most terrestrial vertebrate protein tiles). Notice that the carbon < || oxidized; the organism discarc its available energy of oxidatic eee oe H.N—C—H ¢=0 R E = R Amino acids coo” coo” a-Keto acids | t a-Ketoglutarate Cc=0 N—C—H a vent CH. CH. janine ine ite HN—C—H — from CH. CH. muscle ie [ie CH; Ccoo- coo- a-Ketoglutarate Glutamate = coo” c=0 NH; | CH; Pyruvate _ H,N—C—H | Glutamine not from muscle CHe and b other £ tissues O NBs Glutamine + cyl NHy, urea, or uric acid  Tissular protein Dietary protein Breakdown of peptide bonds Protein synthesis + cofactors Amino acids Amino acid derivatives Deamination Decarboxylation Transamination NH,” Carbon skeletons FA synthesis Fatty acids wo : ¥ Gluconeogenesis Glucose g Ketogenesis Ketone bodies ° TCA cycle ATP Synthesi mus Cholesterol Urea %* Tissular protein breakdown is increased during periods of fasting/starvation Arg Pro His Val De Met Trp Ala Gly Ser Thr Cys Asp Asn Lys Thr Met Ile (Catabolic breakdown of amino acids Anabolic formation of amino acids produces citric acid cycle intermediates uses citric acid cycle intermediates as precursors = FIGURE 23.7 The relationship between amino acid metabolism and the citric acid cycle. Amino Acid Catabolism • The removal of α-amino groups from AA is through: – Transamination by amino transferases (transaminases) in all tissues • This can be after conversion of those AA to Glu – and/or oxidative deamination by glutamate dehydrogenase in hepatocytes • Transamination releases α-keto acids and usually one of the 4 AA mentioned Amino Acid Catabolism • Oxidative deamination releases ammonia and α-ketoglutarate • Some of the free ammonia (in kidney cells) is excreted in urine, but most is captured and channeled into the synthesis of and excreted as urea Amino Acid Catabolism • Urea is quantitatively the most important route for disposing nitrogen from the body • The α-keto acids, AKA ‘carbon skeletons of the deaminated amino acids’, are converted to common intermediates of energy-producing metabolic pathways – AcetylCoA, α-ketoglutarate, succinylCoA, oxaloacetate, fumarate, pyruvate • These compounds can be metabolized to CO2 and water, glucose, fatty acids, and/or ketone bodies by the central pathways of metabolism Transamination • This is the transfer of an amino group from amino acid to an amino group acceptor, usually α-ketoacid producing another set of α-keto acid and amino acid • Aminotransferases (transaminases) catalyze this transfer of amino groups from one carbon skeleton to another Transamination • Transaminases use pyridoxal phosphate (PLP) as a coenzyme – PLP is a coenzyme form of vitamin B6, – PLP accepts an amino group becoming pyridoxamine and passes it to an α-keto acid in a ping-pong reaction style – Transaminases catalyze reversible reactions, are specific to and named after the amino group donor, Transamination • Ultimately the amino groups collected through transamination reactions from other amino acids are transferred to α-ketoglutarate forming glutamate • α-ketoglutarate thus, plays a unique role in AA metabolism by accepting amino groups from other amino acids • The glutamate produced can undergo oxidative deamination in hepatocytes or used as amino group donor in various biosynthetic pathways such as synthesis of non essential amino acids, nucleotide nitrogenous bases in different tissues Reactions catalyzed during amino acid catabolism. ALT was formerly called Glutamate:Pyruvate transaminase (GPT) AST was formerly called Glutamate:Oxaloacetate transaminase (GOT) Aminotransferases • During AA catabolism, in most tissues and especially the liver ALAT catalyzes the transfer of amino group of alanine to α- ketoglutarate, producing pyruvate and glutamate – Skeletal muscles are an exception in this situation, amino groups are transferred to pyruvate and collected as alanine Aminotransferases • During AA catabolism in the liver ASAT catalyzes the transfer of amino group of some glutamates to oxaloacetate to form aspartate and α-ketoglutarate (an exception). – Aspartate supplies the second nitrogen in urea synthesis Diagnostic value of plasma aminotransferases • ALAT and ASAT aminotransferases are usually intracellular enzymes present in many tissues, especially enriched in hepatocytes • The presence of elevated plasma levels of aminotransferases could indicate destruction of tissue cells rich in these enzymes • Plasma ASAT and ALAT are elevated in nearly all liver diseases, particularly in conditions that cause extensive cell necrosis, eg severe viral hepatitis, toxic injury, hypoxic injury etc. Nitrogen Flow in Amino Acid Catabolism a-Amino acid a-Keto acid ama) >< a-Ketoglutarate L-Glutamate OXIDATIVE DEAMINATION NH; co, UREA a Urea  Amino Acid Catabolism • These products directly join central metabolic pathways as intermediates, resulting in lipid or glucose synthesis, or in the production of energy through oxidation by the TCA cycle • Amino acids can be classified as glucogenic, ketogenic or both depending on which of the seven intermediates are produced when they are catabolized Amino Acid Catabolism • Glucogenic amino acids yield pyruvate or one of the TCA cycle intermediates when catabolised. They can give rise to net formation of glucose or glycogen • Ketogenic amino acids yield acetoacetate or one of its precursors (acetylCoA or acetoacetylCoA) upon catabolism • The classification is not absolute, some AA are both ketogenic and glucogenic Glucogenic Vs Ketogenic AAS Alanine | 0rginine* Asparagine Aspartate Cysteine Glutamate Glutamine Ghycine Histidine” Proline Serine nH SN oO ce ll L-Proline L-Glutamate - NAD* | PROLINE DEHYDROGENASE @) a-Ketoglutarate NADH + H* a Ae ll oO H;0 NH3* | Hoye O oO L-Glutamate-y-semialdehyde NAD* GLUTAMATE SEMIALDEHYDE ® DEHYDROGENASE NADH + H* L-Glutamate • Arginase-1 • Cytosol of hepatocytes, macrophages, myeloid cells (inducible) • Arginase-2 • Mitochondrial enzyme in kidney, brain, small intestine, in addition to most cells in the body • Role of arginine in immunity • NO-mediated vasodilation and increased blood supply to a tissue i Ny o Histidine catabolism L-Histidine eHistidinemia NH,* HISTIDASE » Urocanate 0 UROCANASE COOH COOH He=0 HO-NH, ch, cH, CH,COOH du,coon o-Ketoglutaric acid Glutamic acid CH, CH, | 1 HC-NH, a c=0 | alanine aminotransferase | COOH COOH Alanine Pyruvic acid | alll acetyl-CoA CH,-NH, I COOH Glycine serine hydroxmethytransferase NH, H,0 serine dehpdratase CH,OH | HCNH, COOH Serine THF NeN!? methylen-THF THF NSN!-CH,-THF Oxalate <—— Glyoxylate<—— Glycine NH; + CO, OH-Pyr NAD* NADH NH; [€ (13, Désenine NS N!°-CH,-THF H;03 os THF NH; 3-P-serine Pyr —(Co)---> Glucose ; 1 \ a-KG H,0 Pi Pyr » 4 ‘ Phosphatidylethanolamine . Phosphatidylcholine) Glu Ala : \ ( Cysteine | \ : OH-P \ Ethanolamine 3-P-OH-Pyr ree \ (Choline) 1 \ NADH NADH NAD" ® v Phosphatidylserine NAD* Sphingolipids glycerate 3-P-glycerate ,P-elyeer ® co v ATP \ oe Phosphatidylcholine Phosphatidyl- i . ' Glucose | | Pyr 2-P-glycerate Anandamide, ... ethanolamine Pathways of serine synthesis and metabolism •1, serine hydroxymethyltransferase; •2, glycine cleavage system; •3, serine dehydratase; •4, 3-phosphoglycerate dehydrogenase; • 5, phosphoserine aminotrasferase; • 6, phosphoserine phosphatase; • 7, serine-pyruvate aminotransferase; • 8, glycerate dehydrogenase; • 9, glycerate kinase; • 10, phosphatidylserine synthase; • 11, phosphatidylserine decarboxylase; • 12, racemase; • 13, D-amino acid oxidase. AMA Style Holeček M. Serine Metabolism in Health and Disease and as a Conditionally Essential Amino Acid. Nutrients. 2022; 14(9):1987. https://doi.org/10.3390/nu14091987 Cysteine HC [0] a-Keto acid CYSTEINE DIOXYGENASE TRANSAMINASE ca-Amino acid II CH oO _ Re 7 te Cc oO Cysteine sulfinate He ; C Sulfinylpyruva H,C™ oon “O25 oO . DESULFINASE hs sos * Sulfinase pathway of Cysteine catabolism Pyruvate CYSTEINE a-KA TRANSAMINASE a-AA 3-Mercaptopyruvate I o- (thiolpyruvate) HsC~ “cr HS 0 NADH 2H +H* ~=t ww . Pyruvate H2S NAD H <Q -0H . H Cc “C7 3-mercaptopyruvate 2) i pathway of Cysteine HS Oo catabolism 3-Mercaptolactate Oo HO H L | a COCs H NH, Threonine Thr aldolase TDH STDH | Acetatdehyde ) +(_ Glyn wien Castes) [at] + (ee ——_» 1 - Type II tyrosinemia 2 - Neonatal tyrosinemia 3 - Alkaptonuria FUMARYLACETOACETATE 4 - type | tyrosinemia, or tyrosinosis Lysine • Lysine is catabolized in the liver to crotonylCoA • CrotonylCoA is degraded to AcetylCoA and CO2 by reactions of fatty acid degradation pathways • the amino groups are transaminated to α- ketoglutarate to form Glu Branched-chain Amino Acids • Leucine – AcetylCoA and Acetoacetate • Valine – SuccinylCoA • Isoleucine – AcetylCoA and SuccinylCoA Figure 4a, Biotin-containing Carboxylases in the Metabolism of BCAAs, Odd Chain Fatty Acids, and Cholesterol isoleucine leucine valine Bronched-choin onminotronsferase Bronched-chain | lo | dehydrogenase " 4 4 tigly-CoA 3-methyicrotonyl-CoA ——_—methacrylyl-CoA 4 methyfcratonyi-CoA ‘ 3-methyl-3-hydroxybutyryl-CoA corbonyiase = 3 -hydiroxyisobutyryl-CoA 4 3-methylglutaconyl-CoA 4 2-methylacetoacetyl-CoA | methyimalonate 3-hydroxy-3-methylglutaryl-CoA Semiaidehyde Pciicsiad K 4 propionyl-CoA acetoacetate propionyl-CoA propiony!-CoA propionyt-CoA acetyl-CoA - D-malonylmalonyl-CoA D-malonylmalonyl-CoA ‘ ' L-malonyimalonyl-CoA L-malonylmalonyl-CoA + ‘ succinyl-CoA succinyl-CoA \ | I succinyl-CoA t L-malonylmalonyl-CoA t D-malonylmalonyl-CoA ProplonyHteA = acetyl-CoA cholesterol —+ propionyl-CoA «+. -eaigk Branched-chain Amino Acids • The initial reactions are – Transamination – Branched chain amino transferase – Oxidative decarboxylation – Branched chain α-ketoacid dehydrogenase complex • The initial reactions are identical • The transaminase is not found in the liver • Maple syrup urine disease (branched chain ketonuria) Disposal of the Amine/Amide Nitrogen • The effect of transamination and other AA catabolic reactions is to collect the amino groups from many different AAs in the form of L-glutamate • Glutamate then functions as an amino group donor for biosynthetic pathways or for excretion pathways • Ammonia released as NH4+ from other degradative pathways in cells is added to glutamate to form glutamine Disposal of the Amine/Amide Nitrogen • Glu and Gln not used for protein or other biosynthetic reactions is transported to hepatocytes for nitrogen disposal in urea cycle • In cytosol of hepatocytes amino groups of Alanines from skeletal muscles is donated to α-ketoglutarate forming glutamate – These and other glutamates from other sources is transported to the mitochondria Disposal of the Amine/Amide Nitrogen • In mitochondria of hepatocytes the amino group on glutamates is removed regenerating α-ketoglutarate – Oxidative deamination by glutamate dehydrogenase – Transamination by ASAT producing aspartate Transport of ammonia to the liver • Two mechanisms are available in humans for making the non-toxic ammonia carriers • First is the formation of glutamine • The free ammonia produced in tissues is combined with glutamate to yield glutamine by the action of glutamine synthetase Transport of ammonia to the liver • This reaction requires ATP and produces inorganic phosphate • Glutamine is a nontoxic transport form of ammonia; it is normally present in blood in much higher concentrations than other amino acids coo” CH, CH HCNH3* coo” Glutamate ATP + NH, Glutamine synthetase ADP +P, CO -NH, oH CH, HCNH3* coo” Glutamine Figure 19.18 Synthesis of glutamine.  co -NH, oo CHs HCNH,* COO- Glutamine H,0 Glutaminase NH, coo” ee ae HCNH,* CoO” Glutamate Figure 19.17 Hydrolysis of glutamine to form ammonia. Transport of ammonia to the liver • The second mechanism is the transamination of pyruvate into alanine • This is used primarily by skeletal muscles • Therefore alanine also plays a special role in transporting amino groups to the liver in a nontoxic form, via glucose-alanine cycle Transport of ammonia to the liver • The formed alanine passes into the blood and travels to the liver • In the cytosol of hepatocytes, alanine aminotransferase transfers the amino group from alanine to α-ketoglutarate, forming pyruvate and glutamate • In the liver, the pyruvate can be used by the gluconeogenesis pathway to make glucose, hence the alanine-glucose cycle.