AMINO ACID METABOLISM

1. Metabolism of Amino Acids M.Prasad Naidu MSc Medical Biochemistry, Ph.D,.

2. introduction • Proteins  most abundant org.compound • Major part of the body dry wt (10-12Kg) • Perform wide variety of functions. Viz • 1. Static functions ( Structural functions) • 2. Dynamic functions( Enzy, hor, receptors) • Half of the body protein is (Collagen) is present in supportive tissue (skeletan & connective) while the other half is intracellur.

3. introduction • Proteins are the N- containing macro molecules • Consists of L- AAs as repeating units • Of the 20 AAs half can be synthesized • Essential and non-essential AAs • Proteins on degradation release AAs • Each AA undergoes its own metabolism • Proteins metabolism is more appropriately learnt as metabolism of amino acids.

4. Amino Acid Pool • An adult has about 100 gm of Free AA which represent the AA pool of the body. • Glutamate and Glutamine together constitute about 50% and EAA 10% of the body pool. • The conc of intracellular AA is always higher than the Extracellular AA • AAs enter the cells againt Active transport • The AA pool is maintained by the sources that contribute ( input) and the metabolic pathways that utilize (out put) the amino acids.

5. Sources of AA Pool • 1. Turnover of body protein • 2. intake of dietary protein • 3. synthesis of non- EAAs

6. Protein turnover • The protein present in the body is in a dynamic state. • About 300-400 gm of protein per day is constantly degraded and synthesized which represent the body protein turnover. • There is wide variation the turnover of individual proteins. • Eg: plasma proteins & digestive enzymes are rapidly degraded ( half life is hrs/days) • Structural proteins have long half lives often months and years.

7. Control of protein turnover • many factors • 1. Ubiquitin : small PP – 8,500 – tags with the proteins and facilitates degradation. • 2. PEST Sequences: - Certain proteins with Pro, Gln, Ser, Thr sequence are rapidly degraded.

8. Dietary Protein • Regular loss of protein due to degradation of AAs. • About 30-50 gm protein is lost every day from the body. • This amount must be supplied daily in the diet to maintain N Balance. • There is no storage form of AAs unlike the Carbohydrates and lipids (TG) • The excess AAs – metabolised – oxidized –Energy or glucose or fat. • The daily protein intake by adults is 40-100gm

9. Synthesis of AAs • 10 out of 20 naturally occurring AAs can be synthesized by the body which contributes to AA pool.

10. Utilization of AAs from body pool • 1. most of the body proteins (300-400g/D) degraded are synthesized from the AA pool. ( enzymes, hormones, immuno proteins, contractile proteins) • Many imp N compounds ( porphyrins, purines & pyrimidines) are produced from AA . About 30g of protein is daily utilized for this purpose. • Generally, about 10-15% of body energy requirements are met from the AAs • The AAs are converted to Car, fats. This becomes predominant when the protein consumption is in excess of the body requirements.

11. Metabolism of amino acidsgeneral aspects • AAs undergo common reactions • Transamination followed by • Deamination for the liberation of NH3 • The NH2 group of AAs is utilized for the formation of urea (excretory end product of protein metabolism) • The C-skeleton of the AAs is first converted to keto acids (by transamination) which meet one or more of the following fates

12. Fate of keto acids • Utilized to generate energy • Used for the synthesis of glucose • Derived for the formation of fat / ketone bodies • Involved in the production of non-EAAs

13. Transamination • Transfer of an amino group from an AA to a keto acid • This process involves the interconversion of a pair of AAs and a pair of keto acids • Transaminases / aminotransferases

14. Salient features of Transamination • All transaminases require PALP • Specific transaminases exist for each pair of amino and keto acids • However, only two namely Asp. transaminase & Ala. transaminase make a significant contribution for transamination • There is no free NH3 liberated, only the transfer of NH3 group occurs • Reversible • Production of non-EAAs as per the requirement of the cell • Diverts the excess of AAs towards Energy generation

15. Salient features of Transamination • AAs undergo TAN to finally concentrate N in glutamate • Glutamate is the only AA that undergoes OD to liberate free NH3 for urea synthesis • All AAs except Lys, Thr, Pro & Hy.pro participate in TAN • TAN is not restricted to α-group only. (eg: δ-amino group of Ornithine is transaminated. • Serum transaminases are important for diagnostic and prognostic purposes • SGPT or ALT is elevated in all liver diseases • SGOT or AST is increased in myocardial infarction

16. Mechanism of Transamination • Occurs in 2 stages. • 1. Transfer of the NH2 group to the coenzyme PLP ( bound to the coenzyme) to form Pyridoxamine Phosphate. • 2. The NH2 group of Pyridoxamine PO4 is then transferred to a keto acid to produce a new AA and the enzyme with PLP is regenerated.

17. Mechanism of Transamination • All the transaminases require PLP , a derivative of Vit B6 • The – CHO group of PLP is linked with έ-NH2 group of Lys, at the active site of the enzyme forming a Schiff’s base (imine linkage) • When an AA comes in contact with the enzyme, it displaces lys and a new Schiff base linkage is formed. • The AA-PLP-Schiff base tightly binds with the enzyme by non covalent forces. • Snell & Braustein proposed Ping-Pong Bi Bi mechanism involving a series of intermediates ( aldimines & ketimines) in transamination reaction.

18. Deamination • The removal of amino group from the AAs as NH3 • Transamination involves only shuffling of NH3 groups among the AAs • Deamination results in the liberation of NH3 for urea synthesis • Simultaneously, the C-skeleton of AAs is converted to keto acids • 2 types (Oxidative & Non oxidative) • Transamination & Deamination occurs simultaneously, often involving glutamate as the central molecule (Transdeamination)

19. Oxidative deamination • Liberation of free NH3 from the AAs coupled with oxidation • Liver & kidney • Purpose of OD: to provide NH3 for urea synthesis & α-ketoacids for a variety of reactions, including Energy generation

20. Role of GDH • In the process of Transamination, the NH3 groups of most of the AAs are transferred to α-KG to produce glutamate • Thus , glutamate serves as a collection centre for amino groups in the biological system • Glutamate rapidly undergoes oxi.deamination by GDH to liberate NH3 • GDH is unique in that it can use utilize either NAD+ or NADP+ • Conversion of glutamate to α-KG occurs through the formation of α-iminoglutarate • GDH catalyzed reaction is imp as it reversibly links up glutamate metabolism with TCA cycle through α-KG • GDH is involved in both catabolic & anabolic reactions.

21. Regulation of GDH activity • Zn containing mitochondrial enzyme • Complex enzyme containing 6 identical units with a mol.wt of 56000 each. • GDH is controlled by allosteric regulation • GTP , ATP, steroid & Thyroid hormones are inhibitors of GDH • GDP and ADP are activators • After ingestion of protein meal, liver glutamate level is ↑. • It is converted to α-KG with liberation of NH3 • Further , when cellular E levels are ↓low, the degradation of glutamate is ↑ to provide α-KG which enters TCA cycle to liberate Energy

22. Oxidative deamination by AAoxidases • L- AAoxidase & D-AAoxidase are flavo proteins, possessing FMN and FAD respectively. • They act on corresponding AAs to produce α-Ketoacids & NH3 • In this reaction, O2 is reduced to H2O2, which is later decomposed by catalase • The activity of L-AAoxidase is much low while D-AAoxidase is high in tissues (liver & kidneys) • L-AAoxidase does n’t act on Gly & dicarboxylicacids

23. Fate of D-aminoacids • D-AAs are found in plants & mos • Absent in mammalian proteins • But D-AAs are regularly taken in diet and are metabolized • D-AAoxidase converts them into α-ketoacids by od. • The α-ketoacids so produced undergo TAN to be converted to L-AAs • Ketoacids may be oxidized to generate energy or serve as precursor for glucose & fat synthesis • Thus D-AAoxidase is imp as it initiates the first step for the conversion of unnatural D-AAs to L-AAs in the body.

24. Non oxidative deamination • Some of the AAs can be deaminated to liberate NH3 without undergoing oxidation • A) Aminoacid dehydrases: • Ser,Thr,Homoserine α-ketoacids • Catalyzed by PLP dependent dehydrases (dehydratases) • B)Aminoacid desulfhydrases: • Cys, homocysteine  pyruvate • Deamination coupled with desulfhydration • C) Deamination of histidine: • Histidine  urocanate • histidase