Nucleotide MetabolismNucleotide Metabolism

Nucleotide SynthesisNucleotide Synthesis

• De Novo Pathway– Synthesize purine and pyrimidine nucleotides

from low M.W. precursors• Salvage Pathway

– synthesize nucleotides from nucleosides of nucleobases

NB: important targets for therapy of microbial or parasitic diseases

Nucleic Acid DegradationNucleic Acid Degradation

• Intracellular Degradation– Turnover of unstable RNA; cell death; ingested

nucleic acids

• Extracellular Degradation– Major route by which nucleosides or

nucleobases become available in animals

Nucleic Acid DegradationNucleic Acid Degradation

– endonucleases → oligonucleotides

– phosphodiesterases → mononucleotides

– nucleotidases → ortho PO4; nucleosides

– nucleoside phosphorylase → base; ribose-1-P

Nucleoside Nucleoside PhosphorylasePhosphorylase

PRPP is a Central Metabolite in De Novo and Salvage PathwaysPRPP is a Central Metabolite in De Novo and Salvage Pathways

• An intermediate in histidine and tryptophan biosynthesis

• Synthesis of nucleoside-5’-phosphate (nucleotide) from free bases

LowLow––Molecular Weight Precursors to the Molecular Weight Precursors to the PurinePurine RingRing

• Feedback regulation of early steps– PRPP synthetase (–) by AMP, ADP, GDP– PRPP amidotransferase (–) by AMP, GMP

(synergistic inhibition)

Pathways from Pathways from inosinic inosinic acid to GMP and acid to GMP and AMPAMP

• Energy drive is from GTP/ATP– A way to control

the proportion of IMP that go to adenine and guanine nucleotide synthesis

• (reciprocal substrate relation)

Conversion of Nucleoside Conversion of Nucleoside Monophosphate Monophosphate to the to the TriphosphateTriphosphate

• Step 1- Specific ATP-Dependent Kinase

– Guanylate kinase– Adenylate kinase

• Step 2 – Non-Specific ATP-Dependent

– Nuceloside diphosphokinase

Control of Control of Purine Purine BiosynthesisBiosynthesis

• Primates → uric acid• Most mammals further oxidize the purine

ring to allantoin and to allantoic acid or further to urea

Enzymatic abnormalities that lead to Enzymatic abnormalities that lead to hyperuricemia hyperuricemia and gout and gout by elevating the rate of de novo by elevating the rate of de novo purine purine nucleotide biosynthesisnucleotide biosynthesis

GoutGout

• Excessive accumulation of uric acid

• 3/1000 suffer from HYPERURICEMIA– Urate ppt. causes inflammation in the joints→ Painful arthritis

• Allopurinol inhibits xanthine oxidase• Solubility: hypoxanthine/xanthine > uric acid

De Novo Synthesis of De Novo Synthesis of Pyrimidine Pyrimidine NucleotidesNucleotides

• Pyrimidine ring is assembled as a free base

• Unbranched pathway

• Aspartate transcarbamoylase(–) by CTP(+) by ATP

Pyrimidine Pyrimidine RingRing

Control of Control of Pyrimidine Pyrimidine BiosynthesisBiosynthesis

Catabolic Pathways in Catabolic Pathways in Pyrimidine Pyrimidine Nucleotide Nucleotide MetabolismMetabolism

Overview of Overview of Deoxyribonucleotide Deoxyribonucleotide BiosynthesisBiosynthesis

Overview ofOverview of DeoxyribonucleotideDeoxyribonucleotide BiosynthesisBiosynthesis

• Close regulatory relationships between DNA synthesis and dNTP metabolism

• Conversion of ribose to deoxyribose

• Conversion of uracil to thymine

Mechanism for Reduction of a Mechanism for Reduction of a Ribonucleoside Ribonucleoside Diphosphate Diphosphate by by rNDP ReductaserNDP Reductase

• Replacement of the 2’-hydroxyl moiety of the sugar by a hydride ion – retention of configuration

• Ribonucleoside diphosphate reductase (rNDP)

Reductive electron transport sequences in the Reductive electron transport sequences in the action of action of ribonucleoside diphosphate reductaseribonucleoside diphosphate reductase

Ribonucleoside Diphosphate ReductaseRibonucleoside Diphosphate Reductase

• Both activity and specificity being regulated– To maintain balanced pools of DNA precursors

(1) Activity Sites• Low affinity binding of ATP or dATP• ATP binding → (+)• dATP binding → (–)

Ribonucleoside Diphosphate ReductaseRibonucleoside Diphosphate Reductase

(2) Specificity Sites• High affinity binding of ATP, dATP, dGTP or dTTP

• Modulates the activities of enzyme toward different substrates

– Maintain a balanced rate of production of dNTPs

Salvage and De Novo Synthetic Pathways to Salvage and De Novo Synthetic Pathways to Thymine Thymine NucleotidesNucleotides

Relationship between Relationship between thymidylate synthase thymidylate synthase and and enzymes of enzymes of tetrahydrofolate tetrahydrofolate metabolismmetabolism

Thymidylate Thymidylate Synthesis: A Target Site for Synthesis: A Target Site for Cancer ChemotherapyCancer Chemotherapy

• Rapidly dividing cells (eg. cancer cells) require an abundant supply of deoxythymidylate for DNA synthesis

• Thymidylate synthase & Dihydrofolate reductase – TARGET ENZYMES

ThymidylateThymidylate Synthesis: A Target Site for Synthesis: A Target Site for Cancer ChemotherapyCancer Chemotherapy

• Fluorouracil → fluorodeoxyuridylate (F-dUMP)

↓

suicide inhibition of thymidylate synthesis

ThymidylateThymidylate Synthesis: A Target Site for Synthesis: A Target Site for Cancer ChemotherapyCancer Chemotherapy

• Aminopterin and methotrexate inhibit dihydrofolate reductase

• Acute leukemia• Choriocarcinoma

–Rapidly growing tumors