Nucleotide Metabolism Flashcards
1
Q
Roles of nucleotides
A
- Precursors of DNA and RNA
- Universal energy currency(ATP and GTP)
- Components of cofactors
- Molecule activation
- Signal transduction factors
2
Q
Nucleotide Metabolism
A
- De novo synthesis: synthesis of base/sugar from simple precursors.
- Salvage pathway: recycle free bases and nucleosides from polynucleotide breakdown
- Nucleotide degradation: modification of bases to generate products for excretion or reuse
3
Q
Common Intermediates
A
- Pyrimidine -> Uridylate(UMP)
- Purine -> Inosinate(IMP)
4
Q
Similarities between purine and pyrimidine synthesis
A
- Generation of ammonia: Used to source nitrogen group
- Substrate activation by phosphorylation
- Formation of carbon-nitrogen bond
5
Q
Differences between purine and pyrimidine synthesis
A
- Pyrimidine: 4 steps, uses PRPP as ribose-phosphate moiety, PRPP used last, pyrimidine ring synthesized independently, common intermediate is UMP
- Purines: 9 steps, uses 5-ribosyl-1-amine as ribose-phosphate moiety -> used in first step -> purine ring built off it, common intermediate is IMP
6
Q
Pyrimidine synthesis
A
- Bicarbonate + NH3 uses 2 ATP to form carbomyl phosphate -> pyrimidine ring -> PRPP -> UTP to CTP to RNA, TTP to dCTP to DNA
7
Q
Production of Carbomyl Phosphate
A
- Bicarbonate -> Carboxyphosphate -> Carbamic Acid -> Carbomyl Phosphate
8
Q
Carbomyl Phosphate Synthetase
A
- Converts bicarbonate to carbomyl phosphate
- Substrates travel 80A
- Multiple active sites catalyze reactions
- Phosphorylate bicarbonate using ATP
- Hydrolyze NH3 from glutamine
- Add NH3 to carboxyphosphate
- Transfer phosphate from ATP onto carbamic acid to form carbomyl phosphate
9
Q
Aspartate as a substrate: synthesis of open ring
A
- Carbomyl phosphate -> carbamoylaspartate using aspartate substrate and aspartate transcarbamoylase(ATCase)
10
Q
Ring formation
A
- Carbamoylaspartate -> Dihyrooroate -> Oroate
11
Q
Synthesis of UMP
A
- Orotate + PRPP -> orotidylate catalyzed by orotate phosphoribosyltransferase
- Orotidylate -> UMP using orotidylate decarboxylase
12
Q
UTP
A
- Uridylate kinase transfer phosphate from ATP to UMP to form UDP
- Non specific kinase transfer phosphate from any nucleotide to UDP to form UTP
13
Q
CTP
A
- UTP can make CTP by using cytidylate synthetase
- Glutamine ammonia adds amino group to be added to ring -> forms cytosine
14
Q
Pyrimidine de novo synthesis
A
- ATCase joins aspartate to carbamoyl phosphate
- Dihydroorotase closes pyrimidine ring and is oxidized to form oroate
- PRPP used to add ribose 5’-phosphate
- Cytidylate synthetase adds an amino group from Gln to form CTP
- ATP stimulates synthesis while CTP inhibits ATCase
15
Q
Ribonucleotide reductase
A
- Ribonucleotide reductase used to convert NDPs to dNDPs
- combination of R1 and R2 dimers. R1 contains active site with 3 cysteine residues that facilitate catalytic activity. R2 generates free radical to be used in R1 site
16
Q
Reaction Mechanism
A
- Free radical from R2 passes to Cys439 in R1 -> passed to C3’ of bound NDP
- Radical promotes release of water -> formed from the Cys462 H and the C2’ OH
- Cys225 donates H to nucleotide substrate at C2’. Disulfide bond formed between two Cys. Free radical passed from C3’ back to Cys439
- RR reduced by NADPH and TR to return to functional state
17
Q
RR regulation
A
- R1 subunits contain 2 allosteric regulation sites
- Activity site stimulates catalysis by ATP binding, while dATP binding inhibits activity
- Specificity site enhances reduction of pyrimidines when dATP or ATP are bound
18
Q
Synthesis of dTTP
A
- dUDP converted to dUMP by dUTP pyrophosphate
- dUMP used as substrate by thimydylate synthetase
- TS uses THF to source a methyl group -> formation of dTMP
19
Q
Inhibition of thymidylate synthetase
A
- fleurodeoxyuridylate
20
Q
Synthesis of IMP
A
- Start with phosphoribosyl amine
- Addition of glycine
- Addition of carbonyl group
- Addition of amino group
- Closing of 5 membered ring
- Addition of bicarbonate
- Addition of aspartic acid
- Release of fumarate
- Second addition of carbonyl group
- Closing 6 membered ring
21
Q
Purine de novo synthesis
A
- Purine built off ribose sugar
- Amino acids(Asp, Gln, Gly), bicarbonate and THF serve as sources of carbon, nitrogen, oxygen
- NTPs are required for activation and involved in many conversion reactions
22
Q
Functionalization of Purines
A
- Inositate -> Adenylsuccinate using GTP -> AMP using fumarate
- Inositate -> Xanthylate using NAD+ -> GMP using ATP
