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  • 1. Ammonia and amino acid metabolism
  • 2. AMMONIA METABOLISM The ways of ammonia formation1. Oxidative deamination of amino acids2. Deamination of physiologically active amines and nitrogenousbases.3. Absorption of ammonia from intestine (degradation ofproteins by intestinal microorganisms results in the ammoniaformation).4. Hydrolytic deamination of AMP in the brain (enzyme –adenosine deaminase)
  • 3. Ammonia is a toxic substance to plants and animals (especially forbrain)Normal concentration: 25-40 µmol/l (0.4-0.7 mg/l)Ammonia must be removed from the organismTerrestrial vertebrates synthesizeurea (excreted by the kidneys) -ureotelic organismsUrea formation takes place in theliverBirds, reptiles synthesize uric acid
  • 4. Peripheral Tissues Transport Nitrogen to the Liver Two ways of nitrogen transport from peripheral tissues (muscle) to the liver: Glutamate is not1. Alanine cycle. Glutamate is deaminated informed by transamination reactions peripheral tissues
  • 5. Nitrogen is then transferred to pyruvate toform alanine, which is released into the blood.The liver takes up the alanine and converts it backinto pyruvate by transamination.The glutamate formed in the liver is deaminatedand ammonia is utilized in urea cycle.
  • 6. Reaction catalyzed byglutamate dehydrogenase.The glutamate dehydrogenaseof mammalian liver has theunusual capacity to use eitherNAD or NADP as cofactor.The mammalian enzyme isallosterically regulated by GTPand ADP.
  • 7. 2. Nitrogen can betransported as glutamine.Glutamine synthetasecatalyzes the synthesis ofglutamine from glutamate andNH4+ in an ATP-dependentreaction:Ammonia transport in the form ofglutamine.Excess ammonia in tissues is addedto glutamate to form glutamine, aprocess catalyzed by glutaminesynthetase. After transport in thebloodstream, the glutamine entersthe liver and NH4 is liberated inmitochondria by the enzymeglutaminase.
  • 8. Hypotheses toxicity of ammoniaA. The binding of ammonia in the synthesis of glutamate causes an outflow of α-ketoglutarate from the tricarboxylic acid cycle, with decreased formation of ATPenergy and deteriorates the activity of cells.B. Ammonium ions NH4 + caused alkalization of blood plasma. This increases theaffinity of hemoglobin for oxygen (Bohr effect), the hemoglobin does not releaseoxygen to the capillaries, resulting the cells hypoxia occurs.C. The accumulation of free NH4 + ion in the cytosol affects the membranepotential and intracellular enzymes work - it competes with ion pumps, Na + and K+.D. The producing ammonia tramsform glutamic acid - glutamine - an osmoticallyactive substance. This leads to water retention in the cells and the swelling thatcauses swelling of tissues. In the case of nervous tissue it can cause brain swelling,coma and death.E. The use of α-ketoglutarate and glutamate to neutralize the ammonia causes adecrease in the synthesis of γ-aminobutyric acid (GABA) inhibitoryneurotransmitter of the nervous system.
  • 9. THE UREA CYCLEUrea cycle - a cyclic pathway of urea synthesisfirst postulated by H.KrebsThe sources ofnitrogen atoms inurea molecule:- aspartate;- NH4+.Carbon atomcomes from CO2.
  • 10. SYNTHESIS OF UREA Carbamoilphosphate synthase O OHNH 3 + CO 2 + 2 АТФ + H 2O ATP H 2N C O~P O + 2 АДФ + H 3PO 4 ADP OH carbamilphosphate
  • 11. The carbamoyl phosphate generated in themitochondria now donates its carbamoyl group toornithine, which is formed in the cytosol but enters themitochondrion via a spe­cific inner­membrane transportsystem. The product is citruiline: Ornitine-carbomoil transferasa NH 2 NH 2 C O O OH (CH 2)3 NH H 2N C O ~ P O + + H 3PO 4 HC NH 2 (CH 2)3 OH COOH HC NH 2 carbamilphosphate COOH ornitine citrulline
  • 12. The second amino group required for urea synthesisnow arrives in the form of aspartate, which in turnacquired it from glutamate by the action of aspartatetransaminase in the cytosol. The amino group ofaspartate condenses reversibly with the carbamoylcarbon atom of citrulline in the presence of ATP toform argininosuccinate; this reaction is catalyzed byargminosuccinate synthetase:
  • 13. SYNTHESIS OF UREA Arginino-succinate synthase NH2 NH2 COOH C O COOH C N CH NH H2 N CH NH CH2 + + АТФ COOH (CH2)3 CH2 (CH2)3 HC NH2 COOH HC NH2citrulline COOH aspartate COOH NH COOH arginino­succinate C NH CH NH CH2 + АMФ + H4P2O7 COOH (CH2)3 HC NH2 COOH
  • 14. In the next reaction argininosuccinateundergoes a elimination reaction by theaction of argininosuccinate lyase to form freearginine and fumarate: Arginino-succinate lyase NH COOH NH2 COOH C NH CH C NH CH NH CH2 NH + CH (CH2)3 COOH (CH2)3 COOH HC NH2 HC NH2 COOH COOH fumarate arginino­succinate arginine
  • 15. SYNTHESIS OF UREA arginase NH2 NH2 C NH NH2 (CH2)3 NH + H2O + C O HC NH2(CH2)3 NH2 COOHHC NH2 urea COOH ornitinearginine
  • 16. The Linkage between Urea Cycle, Citric Acid Cycle and Transamination of OxaloacetateFumarate formed in urea cycle enters citric acid cycle and is converted to oxaloacetate.Fates of oxaloacetate:(3) transamination to aspartate,(4) conversion into glucose,(5) condensation with acetyl CoA to form citrate,(6) conversion into pyruvate.
  • 17. Diagnostic significance of the determination of urea in urine.25­30 g/day of urea is excreted in normalconditions.The increase of urea in urine occurs inhigh fever, malignant anemia, poisoningby phosphorus, intensive decompositionof protein in organism. The decrease ofurea in urine occurs in liver diseases,kidney unsufficiency, acidosis.
  • 18. Fates of carbon skeleton of amino acids
  • 19. Transformation of AA carbone backbone Serine Glycine Phenylalanine Tyrosine Cystine Homogentizinic acid Cysteine Fumarate Treonine AcetoacetateAlanine Leucine Oxaloacetate SuccinylCoA Pyruvate AcetylCoASerine Ketoglutarate Lysine Glucose Isoleucine Glutamate Urocainic acid
  • 20. epinephrine horepinephrine Triiodtyronine Melanine Dopamine D F Dioxyindol DOPA Tyroxine E A E Phenylalanine Tyrosine Mono­, Diiodtyrosine B B C Phenylpyruvate oxyphenylpyruvate Homogentisinic acid Maleiloacetate Benzoic acid Phenylacetate Phenyllactate FumariloacetateA – PKU D – AlbinosisB – Tyrosinosis, scurvy E – Cretinism, Graves disease Fumarate AcetoacetateC - Alkaptonuria F – Parkinsons’ disease
  • 21. INBORN ERRORS OF AMINO ACIDS METABOLISMAlcaptonuria - inherited disorder of thetyrosine metabolism caused by theabsence of homogentisate oxidase. homogentisic acid is accumulated andexcreted in the urine turns a black color upon exposure to air In children: urine in diaper maydarken In adults: darkening of the ear dark spots on the on thesclera and cornea arthritis
  • 22. Phenylketonuria is caused by an absence or deficiencyof phenylalanine hydroxylase or of itstetrahydrobiopterin cofactor.Phenylalanine accumulates in all body fluids and convertsto phenylpyruvate.Defect in myelination of nervesThe brain weight is below normal.Mental and physical retardations.The life expectancy is drasticallyshortened. Diagnostic criteria:  phenylalanine level in the blood  FeCl3 test  DNA probes (prenatal)
  • 23. NH3 CO2 GlutamateGlutamine Aminobutaric acid methylaspartate TRANSAMINASE carbamylglutamate Pyruvate Acetate ketoglutarate
  • 24. NH3 CO2 ASPARTATEAsparagine Alanine Aspartylphosphate TRANSAMINASE Fumarate Metyonine Lisine Oxaloacetate
  • 25. Maple syrup urine disease - the disorder of theoxidative decarboxylation of α-ketoacids derivedfrom valine, isoleucine, and leucine caused by themissing or defect of branched-chain dehydrogenase.The levels of branched-chain aminoacids and corresponding α-ketoacidsare markedly elevated in both bloodand urine.The urine has the odor of maple syrupThe early symptoms: lethargy ketoacidosis unrecognized disease leads toseizures, coma, and death mental and physical retardation