Nucleotide Metabolism Flashcards
(32 cards)
What is the difference between a nucleotide and a nucleoside?
Nucleotides are phosphate esters of nucleotides.
Nucleoside = nitrogenous base + sugar
Nucleotide = nitrogenous base + sugar + phosphate
Ex/ Nucleotide = Adenine monophosphate, Adenine diphosphate, Adenine Triphosphate (ATP)
What are the two ways (generally) purine nucleotides can be produced?
De Novo synthesis (liver, cytosol) - producing activated sugar first and constructing purine (through 9 step process) onto the ribose sugar Salvage Pathway (organelles) - First making the purine base, then added the ribose sugar to it
What are the two ways (generally) that pyrimidine nucleotides can be produced?
De Novo Synthesis (liver, cytosol, mitochondria) - formation of pyrimidine ring structure followed by addition of ribose phosphate Salvage Pathway (organelles) - formation of pyrimidine nucleotides from pyrimidine bases (RNA/DNA) then adding activated sugar
What are major mechanisms for depletion of amino acid pool?
Protein synthesis (amino acids -> proteins)
Metabolism/degradation of amino acids to ammonia and carbon skeleton
Production of other nitrogen containing compounds (nucleotides)
What are the 4 major steps of purine synthesis?
Phase I: activation of ribose-5-phosphate (from PPP) to make PRPP
Phase II: Convert activated PRPP to even more activated molecule (phosphoribosylamine) —RATE LIMITING
Phase III: Construction of inosine monophosphate (IMP) branch point purine ring
Phase IV: Conversion of IMP into adenosine and guanosine (deoxy) nucleotides
What happens in phase I of purine synthesis?
Activation of ribose 5-phosphate:
Ribose 5-phosphate (from PPP) is phosphorylated to 5-phospho-a-D ribosyl 1-pyrophosphate (PRPP) which is it’s ACTIVE form.
Rxn utilizes ATP -> AMP and enzyme: PRPP synthetase
How is phase I of purine synthesis regulated?
Phase I: Activation of ribose 5-phosphate to 5-phospho-a-D ribosyl 1-pyrophosphate
Enzyme: PRPP synthetase
Requires ATP
Allosterically activated by phosphate levels - Pi levels signal cellular activity due to ATP consumption (high ATP -> activation)
Negatively regulated by levels of purine nucleotides (GMP, AMP, IMP) - feedback inhibition
What happens in Phase II of purine synthesis?
RATE LIMITING STEP
Activated PRPP is converted into phosphoribosylamine (PRA))
(PRPP + Glutamine —> PRA + Glutamate)
Enzyme: Glutamine phosphoribosyl pyrophosphate amidotransferase
Glutamine phosphoribosyl pyrophosphate amidotransferase substitutes pyrophosphate (of PRPP) with amino group (from glutamine) at C-1’ of PRPP. To generate phosphoribosylamine (PRA)
How is Phase II of purine synthesis regulated?
Phase 2: PRPP + Gln —> PRA + Glu RATE LIMITING STEP
Enzyme: glutamine phosphoribosyl pyrophosphate amidotransferase
Allosterically activated by PRPP levels
Negatively regulated by purine nucleotide levels (GMP, AMP, IMP) - Negative feedback inhibition
What happens in phase III of purine synthesis pathway?
PRA enters 9 step ring synthesis to construct IMP (branch point in purine synth)
***Consumes ATP (4 equiv)
2 C’s from formal-THF (folate derivative), 1 C from CO2, remaining C’s and N’s from Gln, Gly, and Asp
What regulates Phase III of purine synth?
Methotrexate (inhibition)
What happens in Phase IV of purine synthesis?
Conversion of IMP to dATP or dGTP
2 pathways:
A Pathway: IMP —> adenylosuccinate monophosphate (requires GTP, adenylosuccinate synthetase) —> AMP (adenylosuccinase) —> ADP —> ATP or dADP —> dATP
G Pathway: IMP —> XMP (*oxidation - requires NAD+ —> NADH; IMP dehydrogenase)
XMP + Gln —> GMP (consumes ATP; releases AMP + PPi and Glu)
GMP —> GDP —> GTP or dGDP—> dGTP
How is Phase IV of purine synthesis regulated?
Negatively Feedback: AMP inhibits rxn of IMP —> adenylosuccinate (adenylosuccinate synthetase)
GMP inhibits rxn of IMP —> XMP (IMP dehydrogenase, ***RATE LIMITING STEP OF GTP SYNTH)
Cross-regulation: AMP synth requires GTP
GMP synth requires ATP
How is purine synthesis regulated?
Feedback inhibition: accumulation of end-product inhibits its own synthesis
AMP/GMP inhibits formation of PRPP, phosphoribosyl amine (PRA), and IMP
Cross-regulation: AMP synth requires GTP/stimulated by GMP
GMP synth requires ATP/stimulated by AMP
(for DNA/RNA synthesis, concentration of end products need to be closely regulated)
What is an anti metabolite?
Therapy targeting nucleotide synthesis
What is Methotrexate? What does it do?
Methotrexate is an antineoplastic agent used for cancer treatment by targeting DNA synthesis in rapidly dividing cancer cells.
- Structurally similar to Dihydrofolate
- Methotrexate COMPETITIVELY INHIBITS dihydrofolate reductase (converts dihydrofolate into bio active tetrahydrofolate) thus cannot form methylene-THF (methyl carrier for rxn of dUMP —> dTMP)
- Methotrexate prevents oxid of NADPH by DHFR
- By reducing formation of dTMP —> reduce DNA synthesis. —> selectively inhibit DNA synth in cancer cells
How does Methotrexate impact purine synthesis?
Phase III of purine synth: 9 step ring assembly requires formyl-THF (f-THF). f-THF comes from m-THF. Therefore methotrexate inhibits conversion of dihydrofolate to tetrahydrofolate —x inhibits production of m-THF —x production of f-THF
What is Fluorouracil? What does it do?
Fluorouracil is a cancer treatment that targets DNA metabolism. Usually used at topical treatment for basal cell carcinoma, warts, injected into tissue for colon, esophageal, breast, cervical cancer.
Specifically targets pyrimidine synthesis.
Targets thymidylate synthase- enzyme catalyzes dUMP —> dTMP (targets same step as methotrexate but different chemical target)
Limits formation of dTMP —X DNA synthesis of cancer cells
What are “sulfa drugs”?
Antibacterial agents in the sulfonamide family that selectively disrupt DNA replication in bacteria
How do Sulfa drugs work?
Bacteria synthesis THF using dihydropteroate synthetase and PABA (p-aminobenzoic acid)
Sulfa drugs mimic the PABA substrate and are competitive inhibitors of dihydropteroate sythetase which halts the THF formation, disrupts nucleotide synthesis and disrupts bacterial DNA synthesis.
What is Acyclovir?
Acyclovir is an antiviral agent that is a synthetic nucleoside analogue
How does Acyclovir work?
Target step: Thymidine kinase phosphorylates the nucleotide deoxythymidine (dT) to generate dTMP using ATP as the phosphoryl donor.
Acyclovir has a base that more closely resembles guanosine than dT (deoxythymidine).
Thymidine kinase preferentially phosphorylates acyclovir, instead of the nucleotide deoxythymidine (dT), for incorporation into DNA synthesis.
Viral TK affinity is much higher for acyclovir than dT. However, Acyclo-dGTP inhibits viral DNA polymerase thus terminating viral DNA replication
Used for treating chicken pox, shingles, and HPV/HSV
Describe purine catabolism
Purine catabolism = breaking down GMP and AMP to Uric acid
GMP dephosphorylated to give –> Guanosine loses ribose group to give —> Guanine is deaminated to give —> Xanthine
AMP de-phosphorylated to give –> Adenosine is deaminated (via adenosine deaminase) to give —> Inosine loses ribose group —> Hypoxanthine is oxidized to give —-> Xanthine
GMP and AMP breakdown are 2 converging pathways that meet at Xanthine
Xanthine is then oxidized (via xanthine oxidase) to give Uric Acid
What does Adenosine Deaminase do? What happens if its defective?
Catalyzes the irreversible deamination of Adenosine to Inosine
Deficiency results in Severe Combined Immunodeficiency (SCID)
Overproduction of erythrocyte isoform causes hemolytic anemia
