Lecture 8: Nucleotide Metabolism Flashcards
What is the difference between nucleosides and nucleotides, and what is a deoxyribonucleotide?
nucleosides: nitrogenous base + sugar
nucleotides: nitrogenous base + sugar + phosphate
deoxyribonucleotide: -H at 2’ C of sugar ring
ribonucleotide: -OH at 2’ C of sugar ring
What are specific roles of nucleotides and nucleosides?
nucleotides: cAMP and cGMP –> regulatory roles, stabilizing regulatory elements (m7GTP cap at 5’ end of eukaryotic mRNA)
nucleosides: in important biomolecules (adenosine in vitamin B12)
What is the difference between de novo synthesis of purine formation vs salvage pathway?
de novo: base formed on ribose 5-phosphate (liver, cytosol)
salvage: ribose-5-phosphate added to preformed base (organelles)
What is the difference between de novo synthesis of pyrimidine formation vs salvage pathway?
de novo: pyrimidine ring followed by ribose phosphate
salvage: pyrimidine nucleotides from pyrimidine bases in RNA/DNA
What are the 4 phases of purine synthesis?
- activation of ribose-5-phosphate
- conversion of PRPP into phosphoribosylamine
- construction of IMP branch point purine ring
- conversion of IMP into adenosine and guanosine (deoxy) nucleotides
Purine Synth Phase I
- ribose-5-phosphate (PPP) –> PRPP
enzyme: PRPP synthetase - (+): phosphate (allosterically); lvls signal cellular activity (ATP consumption)
- (-): purine nucleotides (GMP, AMP, IMP)
Purine Synth Phase II
- PRPP –> phosphoribosylamine (PRA)
enzyme: glutamine:phosphoribonyl pyrophosphate aminotransferase (amino group from glutamine @ C’1) - (+): PRPP
- (-): GMP, AMP, IMP
What is the committed step of Purine Synthesis?
conversion of PRPP into phosphoribosylamine via glutamine:phosphoribosyl pyrophosphate amidotransferase
Purine Synth Phase III
- construction of IMP (9 step ring constructing seq)
- IMP is branch point in anabolism of purines
Methotrexate BLOCKS this process from happening (anticancer drug)
Purine Synth Phase IV
- IMP –> AMP (adenylosuccinate synthetase)
- (-): AMP
- REPLENISH fumarate (TCA cycle)
- IMP –> XMP (NAD–>NADH) –> GMP
- IMP dehydrogenase ((-): GMP)
- GMP synthetase
What are the three phases of pyrimidine synthesis?
- fabrication of pyrimidine ring as OROTATE
- orotate attach to PRPP, generating uridine monophosphate
- uridine monophosphate into cytosine and thymidine nucleotides
Pyrimidine Synth Phase I and Urea Cycle defect
fabrication of pyrimidine ring as OROTATE
- carbamoyl phosphate synthetase
(+): PRPP
(-): UTP - defect in urea cycle = inc. carbamoyl phosphate lvls = Hyperammonemia and Orotic Aciduria
What is the Rate Limiting Step of Pyrimidine Synthesis?
formation of carbamoyl phosphate
Gln –> carbamoyl phosophate –> carbamoyl aspartate
Pyrimidine Synth Phase II
attachment of orotate to PRPP = UMP
UMP synthetase attaches orotate to PRPP, then decarboxylates OMP –> UMP (BIFUNCTIONAL)
What is Orotic Aciduria, where does it occur, and how is it treated?
- inability to convert orotic acid to UMP
- megaloblastic anemia (mental/physical development delays)
- treatment: oral uridine
- occurs in defect of UMP Synthase in Phase II of Pyrimidine Synth
Pyrimidine Synth Phase III
- UMP converted to cytosine and thymidine
cytosine: UTP –> CTP –> dCTP (amination)
thymidine: UMP –> dUDP –> dTTP
- dihydrofolate generation
- thymidylate inhibited by 5-fluorouracil
(dUDP loop used to keep dUTP lvls low so it doesnt get incorporated into DNA)
What are the two key regulatory steps of Pyrimidine Synthesis?
- Carbamoyl phosphate synthetase
(+): PRPP
(-): UMP/UTP - Aspartate transcarbamoylase (ATCase)
(-): CTP
What is the importance of the Pentose Phosphate Pathway?
- produces ribose-5-phosphate and NADPH
- NADPH: reducing environments for glutathione (key antioxidant) –> keeps it from gunking up
liver is principle site of purine/pyrimidine synthesis
What is the importance of Methotrexate?
- cancer treatment
- targets dihydrofolate reductase (DFHR) –> normally converts dietary folate to tetrahydrofolate (binds to it 100x more tightly)
- prevents NADPH oxidation
- disrupts cancer DNA replication
What is the importance of “sulfa” drugs?
- antibacterial agent that competitively inhibits bacterial enzyme incorporating PABA into folate
- DISRUPTS DNA REPLICATION IN BACTERIA
What happens when you deprive cells of GMP and dGTP?
- targeting IMP dehydrogenase
- immunosuppressant disrupting B/T cell DNA rep.
- deprives them of dGTP, helping to prevent rejection
What happens in Purine catabolism?
- ribose removed from guanosine = guanine
- AMP converted to inosine (remove ribose) = hypoxathine
- both create xanthine –> uric acid (GOUT)
Deficiencies in Purine Catabolism (2)
- adenosine deaminase (ADA)
- irreversible deamination (adenosine to inosine)
- overprod: hemolytic anemia
- underprod: SCID
- xanthine oxidase
- leads to uric acid (Gout)
- acid hydrogen, limited water solubility
Severe Combined Immunodeficiency (SCID)
- genetic disorder that compromises B/T cells
- “Bubble Boys” –> need to be isolated from environ.
- ADA deficiency (2nd most common) leads to high levels of adenosine –> dADP/dATP
- this inhibits formation of other dNDPs (impairs DNA synthesis)
What is Gout and how can it be treated?
- high levels of uric acid in blood
Primary and Secondary hyperuricemia
- painful sodium urate deposits in joints (kidneys)
- treatment: allopurinol (inhib xanthine oxidase)
- diet purine diet, alcohol, meat, seafood trigger
Primary vs Secondary Hyperuricemia
Primary = overproduction of uric acid Secondary = underproduction of uric acid
What does Pyrimidine Catabolism produce?
- converts to ketogenic/glucogenic, water-soluble compounds
uracil/cytosine –> malonyl CoA (ketogenic)
thymine –> methylmalony CoA or succinyl CoA (glucogenic)
Purine Salvage
APRT –> adenine to AMP
HGPRT –> guanine/hypoxanthine to GMP/IMP
Lesch-Nyhan Syndrome
- defects in HGPRT (purine salvage), rare form of primary hyperuricemia
- hyperuricemia leads to gout, urate kidney stone, poor muscle control, mental retardation (synth at 200x normal), tendency for self-mutilation
- HGPRT activity <1.5% normal = severe neurological problems, choreoathetosis, self-destructive biting
Kelley-Seegmiller Syndrome
- HGPRT deficiency (>8% normal activity)
- results in Gout, kidney destruction without neurological symptoms
Acyclovir
- antiviral agent that far exceeds viral thymidine kinase phosphorylation (convert to monophosphate dGMP)
- acyclo-dGTP incorporated into rapidly dividing viral cells (terminates DNA replication (no 3’-OH) –> blocks DNA polymerase
- chickenpox, shingles, HPV sore
