Disposal of Nitrogen, Degradation and Synthesis of AminoAcids (2).pptx

df

1. NITROGEN DISPOSAL Nitrogen disposal , Synthesis and Degradation of Amino Acid, Ch 19 , Lippincott’s Dr. Ayman Shahzad

2. Learning Objectives • Understanding the nitrogen metabolism in the human body • Overview of amino acid synthesis and degradation • Exploring the urea cycle and its regulation

3. OVERALL NITROGEN METABOLISM Nitrogen enters the body as D ietary protein. Nitrogen leaves the body as Urea , Ammonia , and other products AMINO ACID POOL All of the FAAs present in cells and ECF throughout the body Supplied by 1) Degradation Of body proteins 2 ) Degradation Of dietary proteins 3 ) Synthesis of non-essential AA from intermediates of metabolism

4. Depleted by S ynthesis of body proteins. S ynthesis of N-containing small molecules purines/pyrimidines/creatinine etc. C onversion of AA into glucose/glycogen/ketone bodies/CO2+H2O PROTEIN TURNOVER The phenomenon of simultaneous synthesis and degradation of proteins . Rate = 300-400g protein being degraded and resynthesized each day

5. PROTEIN DEGARADTION Ubiquitin–proteasome proteolytic pathway: Selective; ATP dependent; Selection of protein to be degraded; C ovalent attachment of protein with UBIQUITIN (4-molecules ) using ATP>> producing a polyubiquitinated protein; recognized by PROTEASOME complex>> unfolds, deubiquitinates, and cuts the target protein into fragments>> fragments then further degraded by cytosolic proteases to AAs

6. PROTEIN DEGRADATION ATP-independent degadative enzyme system of the lysosomes Non-selective; ATP-independent Lysosomes use acid hydrolases to no selectively degrade intracellular proteins (“autophagy”) and extracellular proteins (“heterophagy”), such as plasma proteins, that are taken into the cell by endocytosis

7. Digestion of Dietary Protein by the Proteolytic Enzymes of the Gastrointestinal Tract

8. REMOVAL OF NITROGEN FROM AMINO ACIDS In form of a-amino acids; AA can’t be degraded; remove NH2- group then AA can be metabolized This removal involves Trans- amination Oxidative deamination of amino acids These processes provide aspartate and ammonia for synthesis of urea in urea cycle

9. A. Trans amination: the funneling of amino groups to glutamate Transfer of amino group- via enzymes (aminotransferases). - Requires vitamin B6 (PLP) as a coenzyme . ALANINE AMINO TRASNFERASE ; transfers NH2- from alanine to alpha- ketoglutarate Alanine >> Pyruvate & Alpha- ketoglutarate >> Glutamate ASPARTATE AMINOTRANSFERASE ; Transfers NH2- from Glutmate to OAA Glutamate>> alpha ketoglutarate & OAA>> aspartate ALT and AST used as diagnostic biomarker for liver diseases.

10. B. Oxidative deamination of amino acids Removal of amino group from glutamate via glutamate dehydrogenase, and formation of NH3 and AKA. NH3 liberated exists as NH4+ Glutamate >> Oxidative deamination>> NH3 + A Ketoglutarate Reaction catalyzed by GDH and coenzyme NADH + H+ A Ketoglutarate >> Reductive amination >> glutamate Reaction catalyzed by GDH and coenzyme NADPH + H+

11. C. Transport of ammonia to the liver Two mechanisms; both involving muscles a ) Glutamine formation b ) Alanine formation A) NH3 + Glutamate> GLUTAMINE (in muscle) > glutamine transported to LIVER> where it is cleaved by Glutaminase into glutamate and NH3. NH3 is used in urea synthesis . B) Glucose > pyruvate via glycolysis in muscle ; S uccinyl coA to pyruvate also in muscles; Pyruvate is then converted into ALANINE by transamination catalyzed by ALT; alanine transported out from muscles through blood into LIVER. In liver alanine is converted to pyruvate via transamination by ALT producing glutamate. Glutamate produced in both mechanisms is acted upon by GDH and is converted into AKG and NH3. again NH3 is used for UREA synthesis.

12. Amino Acid Synthesis • Essential vs Non-essential amino acids • Synthesis pathways: - Transamination reactions - Direct synthesis (e.g., glutamate from α- ketoglutarate )

13. Amino Acid Degradation • Ketogenic and Glucogenic amino acids • Examples: - Ketogenic: Leucine, Lysine - Glucogenic: Alanine, Glutam ate, Aspartate - Both: Isoleucine, Phenylalanine Clinical Correlation • Disorders like Phenylketonuria (PKU), Maple Syrup Urine Disease • Role of diet and enzyme replacement therapy

14. UREA CYCLE One nitrogen from free ammonia (by oxidative deamination of Glutamate) and other from aspartate Carbons of urea are derived from HCO3- 1 st 2 reactions occur in mitochondrial matrix while the remaining occur in cytosol 1. Formation of Carbamoyl phosphate: Carbamoyl phosphate synthase I (CPS-I) converts one molecule of NH3 and HCO3- using 2 molecules of ATP to form 1 molecule of carbamoyl phosphate (CP) CPS-I requires N-acetylglutamate as an allosteric activator 2. Formation of Citrulline : Ornithine transcarboxylase (OTC) transfers carbamoyl portion of CP to ornithine to form citrulline. Inorganic phosphate is released Ornithine and citrulline move across mitochondrial membranes via co transporters

15. 3. Synthesis of A rgininosuccinate : Argininosuccinate synthetase combines citrulline with aspartate (containing 2 nd nitrogen of urea) to form argininosuccinate 1 ATP is used and converted into AMP; last ATP of urea cycle; total 3 ATPs are utilized 4. Cleavage of Argininosuccinate : Argininosuccinate lyase cleaves argininosuccinate into fumarate and arginine. Arginine serves as a precursor of urea. Fumarate is hydrated into malate; malate enters mitochondria via malate-aspartate shuttle and is oxidized into OAA by ongoing TCA-cycle in matrix. OAA may be used in gluconeogenesis to form glucose For formation of aspartate by transamination carried out by AST

16. 5. Cleavage of Arginine to Ornithine and Urea : Arginase hydrolyses arginine to ornithine and urea Arginase is present only in liver 6. Fate of Urea : Urea diffuses from liver into blood and is transported to the kidneys where it is excreted. Part of urea is left un excreted and goes to intestine via blood where bacterial UREASE cleaves it into NH3. this ammonia is removed in feces . Kidney failure of urea excretion> more urea in blood> more urea to reach intestine> more bacterial urease activity> more NH3 production> hyperammonemia> may be treated with antibiotics to kill intestinal bacteria containing UREASE. Overall stoichiometry of the urea cycle Aspartate + NH3 + HCO3+ 3 ATP + H2O → urea + fumarate + 2 ADP + AMP + 2 P i + PPi

17. METABOLISM OF AMMONIA A. Sources of ammonia 1. From glutamine: produced from catabolism of branched chain AA; transported via blood to liver kidneys and intestine Liver and kidneys produce NH3 from glutamine by the action of glutaminase and GDH. NH3 produced is converted into urea in case of liver and excreted in case of kidneys, NH3 is excreted as NH4+ in urine therefore plays role in maintaining acid-base balance 2. From bacterial action in the intestine: U rea produced in liver enters blood and reaches kidneys and to some extent intestine. In intestine, bacterial urease converts urea to NH3. This ammonia either enters blood or is removed via feces 3. From amines : 4. From purine s and pyrimidines:

18. B. Transport of ammonia in the circulation Ammonia in its free form can be vey toxic for CNS. Therefore it is transported in blood in form of glutamine or alanine rather than as free NH3 NH3 can be converted into urea for safe disposal via kidneys Or NH3 can be converted into glutamine by combining glutamate and NH3 in the presence of glutamine synthase. Glutamine is a safe transport and storage form of NH3 Glutamine formation occurs mostly in muscles but is of significant importance in CNS to avoid toxic levels of NH3

19. HYPERAMMONEMIA Normal NH3 levels 5-35 micromole/liter Elevated levels due either to liver disease (acquired hyperammonemia) or genetic defects (congenital hyperammonemia) can raise the NH3 blood levels above 1000 micromoles/liter Elevated concentrations of ammonia in the blood cause the symptoms of ammonia intoxication, which include tremors, slurring of speech, somnolence (drowsiness), vomiting, cerebral edema, and blurring of vision . At high concentrations, ammonia can cause coma and death