Chok Biochem 3rd Shift Reviewer Nucleotide Metab

NUCLEOTIDE METABOLISM Purine Nucleotide de novo synthesis Substrate

Product

Enzyme / Coenzyme

Ribose-5-P + ATP

5’-phosphoribosyl-1pyrophosphate (PRPP) + AMP

PRPP synthetase

PRPP + Glu

Β 5-phosphoribosyl amine (β5PRA)

Amidophosphoribosyl transferase (APPT)

Glycinamide ribotide (GAR) FGAR FGAM AIR + ADP + Pi CAIR SAICAR AICAR + Fum FAICAR IMP

GAR synthetase GAR transformylase FGAM synthase AIR synthetase AIR carboxylase (AIRCX) SAICAR synthetase Adenylosuccinate lyase (ASL) AICAR transformylase IMP cyclohydrolase / synthase (IMPCH / IMPS)

Adenylosuccinate + GDP + Pi AMP + Fum

Adenylosuccinate synthetase (ASS) Adenylosuccino lyase

IMP Synthesis β5PRA + Gly GAR FGAR FGAM + ATP AIR + ATP + HCO3GAIR + Asp SAICAR AICAR + N10 formyl THF FAICAR AMP Synthesis IMP + GTP + Asp Adenylosuccinate GMP Synthesis IMP + NAD XMP + Glu + ATP Salvage Pathways Hypoxanthine + PRPP Guanine + PRPP

Xanthosine monophosphate (XMP) + NADH+H GMP + Gln + AMP + PPi IMP + PPi GMP + PPi

Adenine + PRPP

AMP + PPi

adenosine d-adenosine Deoxycytidine D-adenosine 2’d-guanosine

AMP dAMP dCMP dAMP dGMP

IMP DH

Atoms added

Regulation + = Pi, ADP, AMP, GMP, IMP, 2,3bipshophoglycerate - = purine nucleoside di- & triPO4 - = allosteric for IMP/GMP& AMP By Glu, PRPP

Notes Ribose5P from HMP or Ribose1P from nucleoside degradation PRPP α anomer β5PRA β anomer rate limiting / committed / most regulated. irreversible

C4, C5, N7

Reversible

C6

No biotin required

C2, C8

multistep IMP synthesis, uses 6 ATP - = AMP, 6-mercaptopurine

- = GMP, 6-mercaptopurine, tiazopurine

GMP synthetase Hypoxanthine-guanine phosphoribosyltransferase (HGPRT) Adenosyl transferase / Adenine phosphoribosyltransferase (APRT) Adenosine kinase Deoxycytidine kinase

AMP IMP IMP / GMP Inosine / guanosine 5’-nucleotidase Inosine / guanosine Hypoxanthine / guanine Purine nucleoside phosphorylase Guanine Xanthine Hypoxanthine Xanthine Xanthine oxidase - = allopurinol Xanthine Uric acid Xanthine oxidase • Enzyme protein complexes: (1) GAR synthetase + GAR transformylase + AIR synthetase, (2) AIR CX + SAICAR synthetase, (3) AICAR transformylase, IMP CH

Has molybdenum, Fe, sulfur, Final degradation product

Pyrimidine Nucleotide de novo synthesis Substrate

Product

Glu + HCO3- + 2 ATP

Carbamoyl phosphate + Gln + 2 ADP + 2 Pi

Carbamoyl PO4 + Asp

Carbamoyl aspartate

Carbamoyl aspartate DHO + Ubiquinone

Dihydroorotate (DHO) Orotate (Oro) + QH2

Oro + PRPP

OMP + PPi

Enzyme / Coenzyme Carbamoyl phosphate synthase II (CPS II)

Atoms added N3, C2

Aspartate transcabamylase (ATCase)

N1, C4, C5, C6

Regulation + = ATP - = UTP, CTP

Notes Most regulated step Rate limiting step 1st ring ? mitochondrial

Dihydroorotase (DHOase) DHO DH Oro phosphoribosyltransferase (OPRT)

+ = ATP - = UMP, CMP - = UMP, CMP

OMP UMP + CO2 OMP DX UMP + ATP UDP + ADP Nucleoside monoPO4 kinase / UMP kinase UDP + ATP UTP + ADP Nucleoside diPO4 kinase UTP + Glu + ATP CTP + Gln + ADP + Pi CTP synthetase Salvage Pathways Pyrimidine + PRPP Pyrimidine nucleoside monoPO4 + PPi Pyrimidine phosphoribosyltransferase Uracil + Ribose1P Uridine + Pi Uridine phosphorylase Uridine + ATP UMP + ADP Uridine kinase Deoxycytidine + ATP dCMP + ADP Deoxycytidine kinase Thymidine + ATP TMP + ADP Thymidine kinase U UMP U-C kinase C CMP T dTMP T kinase dC dCMP dC kinase Degradation Uracil β-alanine Thymine β-aminoisobutyrate • Enzyme protein complexes: (1) CPS II + ATCase + DHOase, (2) OPRT + OMP DX

- = CTP

Acetyl CoA precursor Succinyl CoA precursor

Comparison Sequence Sources Committed Step Location Enzyme Organization Regulation Deoxyribonucleotide Synthesis Substrate Ribonucleoside DP + thioredoxinH2 Thioredoxin + NADP TMP Synthesis dUMP + N5, N10-methylene tetrahydrofolate dihydrofolate Salvage Pathways Thymine + deoxyribose-1P Thymidine + ATP

Purine Sugar Ring N1 – Asp; C2, 8 – Formate; N3, 9 – Glu; C4, 5, N7 – Gly; C6 – CO2 PRPP amidotransferase / APPT cytosol Individual and 3 multifunctional IMP/AMP/GMP (Product feedback inhibition)

Product deoxyibonucleoside DP + thioredoxin thioredoxinH2 + NADPH+H

Enzyme / Coenzyme Ribonucleoside reductase

tetrahydrofolate

Thymidelate synthetase / N5, N10methylene tetrahydrofolate DHF reductase

Thymidine + Pi dTMP + ADP

Thymine phosphorylase Thymine kinase

dTMP + dihydrofolate

Pyrimidine Ring  Sugar N1, C4-6 – Asp; C2 – CO2; N3 – Glu ATCase (CPS II = most regulated) Mitochondria, cytosol Individual and 2 multifunctional UTP - to CPS

Regulation - = hydroxyurea

Notes Multisubunit enzyme, converts diphosphates into deoxy-forms Regenerates reduced thioredoxin

- = 5-fluorouracil

Contribution of carbon and 2 hydrogen atoms

- = methotrexate

Regenerates reduced thioredoxin

Degradation Step

Substrate

Product

Enzyme

Adenosine deaminase (ADA)

Notes Remove 5’PO4 = -tides to –sides Generally, removes PO4 adenosine  inosine, cytidine  uridine

Nucleoside phosphorylases

Nucleoside + Pi  free bases + ribose1P, reversible

Nucleotidases Hydrolysis Deamination Phosphorylation

Inhibitors Category Antimetabolites Antuifolates Glutamine Antagonists Cell Growth Pur/Pyr analogs

AMP / dAMP Adenosine / deoxyadenosine Inosine + Pi Guanosine + Pi Uridine + Pi Thymidine + Pi

Adenosine / deoxyadenosine Inosine / deoxyinosine Hypoxanthine + Ribose1P Guanine + Ribose1P Uracil + Ribose1P Thymine + Ribose1P

Drug 6-mercaptopurine, 5-fluorouracil, cytosine arabinoside, 6-thioguanine, azathioprine, allopurinol Methotraxate, aminpterin, trimethoprim 6-diazo-5-oxo-L-norleucine (DON), diacetyl-L-serine (azaserine) Hydroxyurea, tiazopurine Acycloguanosine (HSV DNA polymerase), azidothymidine (HIV DNA polymerase)

Diseases • Gout o Precipitation of sodium urate in joints and kidneys due to exceeding serum urate solubility, inflammation and erosion of joints when WBC engulf crystals and rupture o Abdormal / unresponsive PRPP synthetase to feedback inhibition = ↑ purines o Partial deficiency of HGPRT = ↑ PRPP = ↑ purines o Treatment: allopurinol (block urate production) or colchicines – anti-inflammatory, inhibits leukocyte movement • Lesch-Nyhan syndrome o Hereditary X-linked recessive, deficiency of HGPRT (inactive or unstable) = ↑ purine synthesis o Hyperuricemia, gout, urinary tract stones, neurologic symptoms o Treament: allopurinol (↓ urate production) • Von Gierke’s disease o G6PD deficiency --. ↑ Ribose phosphates = ↑ PRPP = ↑ purine synthesis

•

Immunodeficiency disorders o ADA deficiency  ↑ deoxyadenosine and adenosine = ↑ dATP = ↓ ribonucleotide reductase = ↓ DNA synthesis = ↓ lymphocytes  Lymphocyte defect, severe combined immunodeficiency = recurrent, chronic infections, fatal by 18 mo.

 o

Treatment: bone marrow transplant, enzyme replacement, gene therapy Purine nucleoside phosphorylase deficiency  ↑ dGTP = ↓ ribonucleotide reductase to a lesser degree  T-cell impairment = recurrent / chronic infections

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Treament: prevent exposure to diseases