Nucleotide Metabolism Flashcards
1
Q
What are the four purines?
A
- adenosine monophosphate (AMP), DNA/RNA
- guanosine monophosphate (GMP), DNA/RNA
- inosine monophosphate (IMP)
- xanthosine monophosphate (XMP)
2
Q
What are the three pyrimidines?
A
- cytidine monophosphate (CMP), DNA/RNA
- uridine monophosphate (UMP), RNA
- thymidine monophosphate (TMP), DNA
3
Q
What are the roles of nucleosides and nucleotides outside of being building blocks for DNA and RNA synthesis?
A
- nucleotides (e.g. ATP, GTP) are cosubstrates in an array of enzymatic reactions
- nucleotides are components of several cofactors, including CoA, FAD, FMN, UDP-Glc, NADPH, and NADH
- nucleotides (e.g. cAMP and cGMP) play regulatory roles are serve as stabilizing elements, such as m7GTP cap at the 5’ end of eukaryotic mRNA
- nucleosides are in important biomolecules, such as adenosine in vitamin B12
4
Q
What are the sources for pyrimidine bases? (3)
A
- glutamine amide
- aspartate
- bicarbonate
5
Q
What are the sources for purine bases? (5)
A
- glutamine amide
- aspartate amine
- bicarbonate
- glycine
- formate
6
Q
What are the 4 phases of purine synthesis?
A
- activation of ribose 5-phosphate
- conversion of PRPP into phosphoribosylamine
- construction of inosine monophosphate branch point purine ring
- conversion of IMP into adenosine and guanosine (deoxy) nucleotides
7
Q
What happens during phase I of purine synthesis?
A
activation of ribose 5-phosphate
- ribose 5-phosphate (prod of PPP) is converted to “active” form, 5-phosphate-α-D ribosyl 1-pyrophosphate (PRPP) (utilizes ATP and requires PRPP synthetase)
- activated by phosphate levels; Pi levels signal cellular activity due to ATP consumption
- inhibited by levels of purine nucleotides: GMP, AMP, IMP
8
Q
What happens during phase II of purine synthesis?
A
conversion of PRPP into phosphoribosylamine
- glutamine:phosphoribosyl pyrophosphate amidotransferase subs pyrophosphate with an amino group at PRPP C1 and generates phosphoribosylamine (PRA) (amino group is from glutamine)
- activated by PRPP
- inhibited by purine nucleotides
9
Q
What happens during phase III of purine synthesis?
A
construction of IMP
- branch point purine ring
- PRA enters a nine-step ring-constructing sequence that produces IMP
- all intermediates are phosphorylated (nucleotides) due to phosphate goup on ribose 5-phosphate
- consumes 4 ATP
- constituents: folate derivative, CO2, Gln, Gly, Asp
10
Q
What happens during phase IV of purine synthesis?
A
conversion of IMP into dATP and dGTP
- adenylosuccinate synthetase and IMP dehydrogenase inhibited by AMP and GMP respectively
- AS synthetase converts IMP to adenylosuccinate monophosphate, which is converted to AMP and fumarate by adenylosuccinase, AMP converted to ADP, ADP converted to dADP by ribonucleotide reductase, and finally converted to dAMP
- IMP dehydrogenase (requires NAD+) converts IMP to XMP (rate limiting), GMP synthase converts XMP to GMP, GMP converted to GDP, GDP converted to dGDP by ribonucleotide reductase, and finally dGDP converted to dGMP
11
Q
How is purine synthesis regulated?
A
- feedback inhibition: accumulation of end product inhibits its own synthesis
- cross regulation: AMP synthesis is stimulated by GTP, GMP synthesis is stimulated by ATP
12
Q
What are the 3 phases of pyrimidine synthesis?
A
- fabrication of pyrimidine ring as orotate
- attachment of orotate to PRPP to generate uridine monophosphate, the branch point pyrimidine ring
- conversion of uridine monophosphate into cytosine and thymidine (deoxy) nucleotides
13
Q
What happens during phase I of pyrimidine synthesis?
A
fabrication of pyrimidine ring as orotate
- rate limiting step: formation of carbamoyl phosphate from Gln by CPS II
- CPS II activated by PRPP and inhibited by UTP
- urea cycle defect can result in elevated levels of carbamoyl phosphate and manifest as hyperammonemia w/ orotic aciduria
14
Q
What happens during phase II of pyrimidine synthesis?
A
attachment of orotate to PRPP to generate UMP
- UMP synthetase attaches orotate to PRPP to give orotidine monophosphate (OMP)
- UMP synthetase decarboxylates OMP to generate UMP
15
Q
What happens during phase III of pyrimidine synthesis?
A
conversion of UMP into cytosine and thmidine (deoxy) nucleotides
- dUDP loop is wasteful, but thought to occur so that dUTPase keeps dUTP low to prevent incorporating it into DNA
- dUMP is bridge to thymidine production > dTMP > dTDP > dTTP
- UTP is aminated to form CTP > CDP > dCDP > dCMP
16
Q
How is pyrimidine synthesis regulated?
A
- carbamoyl phosphate synthetase: inhibited by UMP/UTP; activated by PRPP
- aspartate transcarbamoylase (ATCase): inhibited by CTP
17
Q
- produces ribose 5-phosphate and NADPH
- in erythrocytes, occurs in liver, testes, mammary glands, and adrenal cortex
- uses NADPH to maintain reducing environments (reduced form of glutathione) and to provide reducing power for biosyn of FA’s and steroids
- formation of glutathione (GSH) is a key antioxidant in detox and reducing oxidative damage
- in humans, liver is principle site of purine and pyrimidine syn and utilizes ribose 5-phosphate and 3/5 principle free AA’s in the liver (Asp, Gln, Gly) as starting material
A
pentose phosphate pathway (PPP)
18
Q
- antineoplastic agent used to treat cancer
- targets dihydrofolate reductase (DHFR), which converts dietary folate to the biologically active tetrahydrofolate in the liver
- prevents oxidation of NADPH
- inhibition disrupts DNA replication in rapidly dividing cancer cells
- efficacy depends on selective drug uptake by cancer cells compared to normal cells
A
methotrexate
19
Q
- antibacterial agents in the sulfonanilamide family that competitively inhibit bacterial enzyme that incorporates p-aminobenzoic acid (PABA) into folate
- PABA is an intermediate in the syn of folate by bacteria, plants, and fungi
- these drugs selectively disrupt DNA replication in bacteria
- humans acquire folate as a vitamin in their diets, thus we are not affected and are selective to bacteria
A
sulfa drugs (antibiotics)
20
Q
- oxidative step in conversion of IMP to XMP is an opportunity for therapeutic intervention
- IMP dehydrogenase is target
- therapeutic immunosuppressant disrupts DNA replication in B and T cells
- deprives them of adequate supplies of GMP, and hence, dGTP
- disrupts GMP synthesis and is useful in preventing graft rejection
A
depriving cells of GMP and dGTP
21
Q
What are the general steps of purine catabolism?
A
- ribose removed from guanosine and inosine to produce gianine and hypoxanthine respectively
- these are both converted to xanthine
- xanthine largely converted to uric acid
22
Q
What are the 2 most important enzymes in purine catabolism?
A
- adenosine deaminase (ADA): irreversible hydrolytic deamination of adenosine to inosine; overprod of erythrocyte isoform causes hemolytic anemia; underprod a/w SCID
- xanthine oxidase: converts hypoxanthine to xanthine and xanthine to uric acid; target for gout treatment
23
Q
What is the relation of oxidation with purine catabolism?
A
- typically, catabolic processes include steps where oxygen is added to make them more polar
- adenine = 0 ox, guanine/hypoxanthine = 1 ox, xanthine = 2 ox, uric acid = 3 ox
- uric acid is the end point of purine catabolism and is the most oxidized
- has an acid hydrogen and limited solubility in water that plays a key role in gout
24
Q
- fatal genetic disorder in which both arms (T and B cells) of adaptive immune system are compromised
- often males bc most common form is x-linked
- early onset failure to thrive, chronic diarrhea, thrush, recurrent infections
- mutations to receptors involved shared by IL’s involved in development and differentiation of T and B cells
- ADA deficiency is the second most common, autosomal recessive inheritance
- leads to increase in adenosine and decrease in inosine
- adenosine converted to AMP and ADP, then to dADP and dATP
- increase in dATP inhibits activity site of ribonucleotide reductases that in turn blocks formation of all other dNDPs
- decrease in dNDPs and dNTP impairs DNA syn and leads to compromised immune system
A
severe combined immunodeficiency (SCID)
25
What are the uric acid markers for gout?
- adult males: 4-8.6 mg/dL, females: 3-5.9 mg/dL
- urinary urate levels are normally \<750 mg/24 hr
- serum urate levels \>9 mg/dL increase the risk of gout
- urinary urate levels are variable day to day, most reliable when patients are on a low purine diet
26
- high levels of uric acid in the blood
- primary hyperuricemia \> overprod of uric acid
- secondary hyperuricemia \> underexcretion of uric acid
- extremely painful deposits of sodium urate in joints of extremities, 3+ million cases each year
- triggers: diets rich in purines (beans, spinach, lentils), alcohol, meat, seafood
- acidemia stimulates reabsorption in the kidney by URAT1
- tx: reducing amnt of granulocytes to affected areas, allopurinol PO that inhibits xanthine oxidase and increases levels of more soluble purines hypoxanthine/guanine
gout
27
What are the general steps of pyrimidine catabolism?
- converted to readily metabolized ketogenic or glucogenic, water-soluble compounds
- end products are malonyl CoA (ketogenic), methymalonyl CoA (glucogenic), and succinyl CoA (glucogenic)
- uracil/cytosine \> malonyl CoA
- thymine \> methylmalonyl CoA or succinyl CoA
28
What is the general mechanism of salvage pathways for nucleotide synthesis?
- bases recovered during nucleotide turnover or digestion can be reincorporated into nucleotides
- salvage dominates over de novo syn for purines
- adenine phosphoribosyltransferase (APRT) generates AMP
- hypoxanthine-guanine phosphoribosyltransferase (HGPRT) generates GMP or IMP
29
- results from defects in HGPRT in purine salvage pathway and is a rare form of primary hyperuricemia
- sx: gout, kidney stones, poor muscle control, mental retardation, self-mutilation
- normally, IMP and GMP are maintained by de novo syn from PRPP and salvage pathway by HGPRT
- excess purines are processed by nucleotidases and nucleosidases that convert GMP and IMP to guanosine and inosine, and ultimately to uric acid
- excess guanine and hypoxanthine shunted to form 6x normal levels of uric acid
- purine biosyn proceeds at levels 200x normal, causes the mental retardation and self-mutilation
- RPP which is not used in salvage pathway is available for additional purine biosynthesis and allosterically activates the next enzyme in purine biosynthesis (Phase II: Glutamine:phosphoribosyl pyrophosphate amidotransferase that generates PRA). Additional PRPP leading to more PRA has mass action effect on additional synthesis for more purines
Lesch-Nyhan syndrome
30
What is the role of HGPRT function in determining LNS severity?
- \<1.5%: LNS presents with additional severe neurologic problems, including spastic cerebral palsy, choreoathetosis and self-destructive biting (fingers and lips)
- \>8%: Kelley-Seegmiller syndrome results with gout and kidney destruction without neurologic symptoms
- 8-15%: variant LNS with neurological problems, ranging from clumsiness to motor dysfunction. Allopurinol can reduce joint and kidney problems but has no effect on neurologic ones
31
- thymidine kinase phosphorylates nucleotide deoxythymidine (dT) to generate dTMP using ATP as phospho donor
- this medication (has a base that more closely resembles guanine than dT) undergoes phospho by viral thymidine kinase at a rate that far exceeds that of cellular kinase
- viral thymidine kinase rapidly converts this med to its monophosphate acyclo-dGMP
- other kinases then convert acyclo-dGMP to acyclo-dGTP
- acyclo-dGTP lacks a 3'-OH group which terminates DNA replication
- this medication is used to help heal sores related to chicken pox, shingles, and HSV
acyclovir
