1. Metabolism of purines and pyrimidines Vladimíra Kvasnicová The figure was found at http://www.mahidol.ac.th/mahidol/ra/rapa/mong/26uric.jpg (Jan 2008) http://www.mahidol.ac.th/mahidol/ra/rapa/mong/26uric.jpg

2. The figure was adopted from Devlin, T. M. (editor): Textbook of Biochemistry with Clinical Correlations, 4th ed. Wiley ‑ Liss, Inc., New York, 1997. ISBN 0 ‑ 471 ‑ 15451 ‑ 2 PURINE BASES

3. The figure was adopted from Devlin, T. M. (editor): Textbook of Biochemistry with Clinical Correlations, 4th ed. Wiley ‑ Liss, Inc., New York, 1997. ISBN 0 ‑ 471 ‑ 15451 ‑ 2 PYRIMIDINE BASES

4. Structure of purine and pyrimidine nucleotides nucleotide = ester of phosphoric acid and a nucleoside nucleoside = N-containing base + monosaccharide  -N-glycosidic bond between base and saccharide nucleotide bases: aromatic heterocycles  purines: pyrimidine + imidazol ring  pyrimidines: pyrimidine ring

5. The figure was adopted from Devlin, T. M. (editor): Textbook of Biochemistry with Clinical Correlations, 4th ed. Wiley ‑ Liss, Inc., New York, 1997. ISBN 0 ‑ 471 ‑ 15451 ‑ 2 ribonucleoside deoxyribonucleoside N-glycosidic bond

6. The figure was adopted from Devlin, T. M. (editor): Textbook of Biochemistry with Clinical Correlations, 4th ed. Wiley ‑ Liss, Inc., New York, 1997. ISBN 0 ‑ 471 ‑ 15451 ‑ 2 ribonucleosides deoxyribonucleoside

7. The figure was adopted from Devlin, T. M. (editor): Textbook of Biochemistry with Clinical Correlations, 4th ed. Wiley ‑ Liss, Inc., New York, 1997. ISBN 0 ‑ 471 ‑ 15451 ‑ 2 Ribonucleotides * N-glycosidic bond * ester bond * anhydride bond

8. The figure was adopted from Devlin, T. M. (editor): Textbook of Biochemistry with Clinical Correlations, 4th ed. Wiley ‑ Liss, Inc., New York, 1997. ISBN 0 ‑ 471 ‑ 15451 ‑ 2 ribonucleotide deoxyribonucleotide

9. Classification of nucleotides purine nucleotides: contain adenine, guanine, hypoxanhine or xanthine pyrimidine nucleotides: contain cytosine, uracil or thymine ribonucleotides (saccharide = ribose) deoxyribonukleotidy (saccharide = deoxyribose)  formed by reduction of ribonucleoside diphosphates (NADPH)

10. The figure was found at http://web.indstate.edu/thcme/mwking/amino-acid-metabolism.html (Jan 2007)http://web.indstate.edu/thcme/mwking/amino-acid-metabolism.html 3´-phosphoadenosine-5´-phosphosulfate (PAPS) used as the sulfate donor in metabolic reactions (sulfatation)

11. Properties of nucleotides strong absorption of UV radiation (260 nm) purines are less stable under acidic conditions than pyrimidines polar terminal phosphate groups  alternative names: adenylate or adenylic acid,...

12. Purine and pyrimidine nucleotides essential for all cells mainly 5´-nucleosidedi and triphosphates ribonucleotides: concentration of a sum of them is constant (mM), only their ratio varies (main ribonucleotide of cells: ATP) deoxyribonucleotides: their concentration depends on a cell cycle (µM)

13. Nucleotides in a metabolism 1) energetic metabolism ATP = principal form of chemical energy available to cells – „as money of the cell“ (30 kJ/mol / spliting off phosphate)  phosphotransferase reactions (kinases)  muscle contraction, active transport 2) monomeric units of RNA and DNA  substrates: nucleoside triphosphates

14. The figure was adopted from Harper´s Illustrated Biochemistry 26 th ed./ R.K.Murray; McGraw-Hill Companies, 2003, ISBN 0-07-138901-6. Synthetic analogs of purines and pyrimidines are used in chemotherapy

15. The figure was found at http://www.benbest.com/health/cycAMP.gif (Jan 2008) http://www.benbest.com/health/cycAMP.gif Cyclic adenosine monophosphate (cAMP) 3) physiological mediators cAMP, cGMP („second messengers“)

16. 4) components of coenzymes  NAD +, NADP +, FAD, CoA The figures were found at http://lxyang.myweb.uga.edu/bcmb8010/pic/NAD+.gif a http://oregonstate.edu/instruct/bb450/stryer/ch14/Slide26.jpg (Jan 2008)http://lxyang.myweb.uga.edu/bcmb8010/pic/NAD+.gif http://oregonstate.edu/instruct/bb450/stryer/ch14/Slide26.jpg

17. 5) activated intermediates  UDP-Glc, GDP-Man, CMP-NANA  CDP-choline, ethanolamine, diacylglycerol  SAM  methylation  PAPS  sulfatation 6) allosteric efectors  regulation of key enzymes of metabolic pathways

18. PRDP = 5-fosforibosyl-1-diphosphate The figure was found at http://ead.univ- angers.fr/~jaspard/Page2/COURS/2N2NH3aaetUree/2Figures/9AAaromatiques/8PRPP.gif (Jan 2008)http://ead.univ- angers.fr/~jaspard/Page2/COURS/2N2NH3aaetUree/2Figures/9AAaromatiques/8PRPP.gif = the substrate for synthesis of both purines and pyrimidines

19. PRPP = 5-phosphoribosyl-1-diphosphate its synthesis is a key reaction of synthesis of the nucleotides PRPP-synthetase is regulated by feed back inhibition by nucleoside di- and triphosphates precursors:* ribose-5-phosphate (from HMPP) * ribose-1-phosphate (phosphorolysis of nucleosides)

20. function:  regulation of nucleotide synthesis  substrate of nucleotide synthesis The figure was adopted from Devlin, T. M. (editor): Textbook of Biochemistry with Clinical Correlations, 4th ed. Wiley ‑ Liss, Inc., New York, 1997. ISBN 0 ‑ 471 ‑ 15451 ‑ 2 PRPP = PRDP

21. Synthesis of purine nucleotides de novo = new building of a nucleotide rings salvage reactions = synthesis from bases or nucleosides  less energy need than for de novo synthesis  they inhibit de novo synthesis  substrates: a) base (adenine, guanine, hypoxanthine) PRPP b) ribonucleosides ATP

22. The figure was found at http://web.indstate.edu/thcme/mwking/nucleotide-metabolism.html (Jan 2007)http://web.indstate.edu/thcme/mwking/nucleotide-metabolism.html Synthesis of purine nucleotides CYTOPLASMCYTOPLASM

23. The figure was adopted from Devlin, T. M. (editor): Textbook of Biochemistry with Clinical Correlations, 4th ed. Wiley ‑ Liss, Inc., New York, 1997. ISBN 0 ‑ 471 ‑ 15451 ‑ 2 AMP GMP IMP

24. The figure was adopted from Devlin, T. M. (editor): Textbook of Biochemistry with Clinical Correlations, 4th ed. Wiley ‑ Liss, Inc., New York, 1997. ISBN 0 ‑ 471 ‑ 15451 ‑ 2

25. The figure was found at http://www.dentistry.leeds.ac.uk/biochem/MBWeb/mb2/part1/aacarbon.htm (Jan 2008)http://www.dentistry.leeds.ac.uk/biochem/MBWeb/mb2/part1/aacarbon.htm Bacteria can synthesize the folate: sulfonamides are analogs of PABA → bacteriostatic effect Folate is a vitamin – it is not synthesized in human cells

26. The figure was found at http://oregonstate.edu/instruct/bb450/lecturenoteskevin/enzymesoutline.html (Jan 2008)http://oregonstate.edu/instruct/bb450/lecturenoteskevin/enzymesoutline.html cytostatics

27. The figure was found at http://www.dentistry.leeds.ac.uk/biochem/postgrad/fol-red.gif (Jan 2008)http://www.dentistry.leeds.ac.uk/biochem/postgrad/fol-red.gif Activation of folate (reduction) Dihydrofolate reductase can be inhibited by Methotrexate

28. The figure was found at http://www.dentistry.leeds.ac.uk/biochem/postgrad/thftypes.gif (Jan 2008)http://www.dentistry.leeds.ac.uk/biochem/postgrad/thftypes.gif Derivatives of tetrahydrofolate

29. The figure was found at http://www.med.unibs.it/~marchesi/glycine_synth.gif (Jan 2008)http://www.med.unibs.it/~marchesi/glycine_synth.gif Serin is the principal donor of methylene group

30. The figure was found at http://www.prema-eu.org/folatepathway/fig1.gif (Jan 2008)http://www.prema-eu.org/folatepathway/fig1.gif Folate in a metabolism

31. Synthesis of purine nucleotides de novo (I) high consumption of energy (ATP) cytoplasm of many cells, mainly in the liver substrates:* 5-phosphoribosyl-1-diphosphate (= PRDP = PRPP) * amino acids (Gln, Gly, Asp) * tetrahydrofolate derivatives, CO 2 coenzymes: * tetrahydrofolate (= THF) * NAD +

32. important intermediates:  5´-phosphoribosylamine  inosine monophosphate (IMP) products: nucleoside monophosphates (AMP, GMP) interconversion of purine nucleotides:  via IMP = common precursor of AMP and GMP (inosine monophosphate: base = hypoxanthine) Synthesis of purine nucleotides de novo (II)

33. Synthesis of pyrimidine nucleotides de novo = new building of a nucleotide rings salvage reactions = synthesis from bases or nucleosides  substrates: a) * base (not cytosine) * PRPP b) * ribonucleosides * ATP

34. The figure was found at http://web.indstate.edu/thcme/mwking/nucleotide-metabolism.html (Jan 2007)http://web.indstate.edu/thcme/mwking/nucleotide-metabolism.html Synthesis of pyrimidine nucleotides CYTOPLASMCYTOPLASM mitochondrion

35. The figure was adopted from Devlin, T. M. (editor): Textbook of Biochemistry with Clinical Correlations, 4th ed. Wiley ‑ Liss, Inc., New York, 1997. ISBN 0 ‑ 471 ‑ 15451 ‑ 2

36. Synthesis of thymidine monophosphate

37. Synthesis of pyrimidine nucleotides de novo (I) cytoplasm of cells (exception: one enzyme is found at mitochondria /dihydroorotate-DH) substrates:* carbamoyl phosphate (Gln,CO 2,2ATP) * aspartate * PRPP * methylene-THF (only for thimidine) Karbamoyl phosphate is formed in urea synthesis as well (only in mitochondria of hepatocytes)

38. important intermediates: * orotic acid (pyrimidine derivative) * orotidine monophosphate (OMP) * uridine monophosphate (UMP) products:* cytidine triphosphate ( from UTP) * deoxythimidine monophosphate ( from dUMP) Synthesis of pyrimidine nucleotides de novo (II)

39. The figure was found at http://www.chm.bris.ac.uk/motm/vitaminb12/ribred.gif (Jan 2008)http://www.chm.bris.ac.uk/motm/vitaminb12/ribred.gif Synthesis of 2-deoxyribonucleotides The reaction is catalyzed by ribonucleotide reductase NADP + NADPH+H + protein

40. Regulation of nucleotide synthesis PRPP-synthetase is inhibited by both purine and pyrimidine nucleoside di- and triphosphates nucleotide synthesis: feed back inhibition nucleoside diphosphate reductase: activated by nucleoside triphosphates, inhibited by deoxyadenosine triphosphate (dATP)

41. The figure was found at http://www.med.unibs.it/~marchesi/purine_synth_reg.gif (Jan 2008)http://www.med.unibs.it/~marchesi/purine_synth_reg.gif Regulation of synthesis of purine nucleotides

42. The figure was found at http://www.med.unibs.it/~marchesi/pyrimidine_synth_reg.gif (Jan 2008) http://www.med.unibs.it/~marchesi/pyrimidine_synth_reg.gif Regulation of synthesis of pyrimidine nucleotides

43. Regulation of nucleotide synthesis regulatory enzymeactivationinhibition glutamine-PRPP amidotransferase (purines)  PRPP  IMP, GMP, AMP (allosteric inhibition) carbamoylphosphate synthetase II = cytosolic (pyrimidines)  PRPP  ATP  UTP

44. Degradation of purines and pyrimidines exogenous: mostly not used for resynthesis endogenous:  enzymes * nucleases (split off nucleic acids) * nucleotidases (...nucleotides) * nucleoside phosphorylases (nucleosides) * deaminase (adenosine) * xanthinoxidase (substrates: hypoxanthine, xanthine) inhibited by allopurinol (pharmacology)

45. The figure was found at http://www.med.unibs.it/~marchesi/purine_degradation.gif (Jan 2008) http://www.med.unibs.it/~marchesi/purine_degradation.gif Degradation of purines

46. URIC ACID keto and enol form salts of uric acid = urates pH of blood: mononatrium urate

47. The figure was adopted from Color Atlas of Biochemistry / J. Koolman, K.H.Röhm. Thieme 1996. ISBN 0-86577-584-2

48. Degradation of pyrimidines

49. SUMMARY:  purines → NH 3, uric acid – it has antioxidative properties (partially excreted with urine; failure: hyperuricemia, gout) physiological range: serum220 – 420 µmol/l (men) 140 – 340 µmol/l (women) urine0,48 – 5,95 mmol/l  pyrimidines: C, U →  -alanine, CO 2, NH 3 T →  -aminoisobutyrate, CO 2, NH 3 The figures were adopted from http://www.uni-koeln.de/med-fak/biochemie/biomed/versuche/v07/abb05.gif and http://www.healerpatch.com/images/gout.jpg (Jan 2008)http://www.uni-koeln.de/med-fak/biochemie/biomed/versuche/v07/abb05.gif http://www.healerpatch.com/images/gout.jpg free radicals

50. Principal differences between metabolism of purines and pyrimidines purinespyrimidines formation of N-glycosidic bond in 1 st step of their biosynthesis (PRDP is the 1 st substrate) a heterocyclic ring is formed first, then it reacts with PRDP location of biosynthesis cytoplasmcytoplasm + 1 enzyme is in a mitochondrion products of degradation uric acid (poor solubility in H 2 O), NH 3 CO 2, NH 3,  -AMK (soluble in H 2 O)