NUCLEOTIDES LECTURE 3

Question 1

why is DNA the genetic material?

Answer 1

removing the 2’ OH makes the DNA phosphodiester backbone more stable
the base thymine allows the detection of spontaneous deamination of cytosine
cytosine, when deaminated, turns into uracil

Question 2

relative oxidation level of DNA and RNA?

Answer 2

DNA is more reduced

Question 3

at which level is the synthesis of deoxyribonucleotides done?

Answer 3

done at the DNPs levels (diphosphates)

Question 4

what are the ratios of NADPH/NADP+ and NAD+/NADH?

Answer 4

Question 5

which enzyme does that work?

Answer 5

ribonucleotide reductase (RR)

Question 6

where do the reducing equivalents originate for RR and where to they pass to?

Answer 6

reducing equivalents originate on NADPH
pass through thiols (on cysteines) of thioredoxin reductase to thioredoxin and finally to RR

Question 7

characteristics of RR?

Answer 7

• dimer of dimers
• reg. and catalytic subunit
• two allosteric sites
• controlled by ATP/dATP levels
• specificity controlled by dATP, dGTP, dTTP
• unusual stable enzyme tyrosine free radical
• inhibited by hydroxyurea
• therapeutic target
• other types exist with other cofactors such as B12, Fe/S

Question 8

what does incorporation of uracil into DNA induce?

Answer 8

induces base excision repair:
1. uracil DNA glycosylase removes uracil base
2. AP endonuclease cleaves DNA backbone
3. resynthesis from complementary strand
4. excessive repair causes mutations

Question 9

what are the starting materials in the pyrimidine de novo pathway and what is the final goal molecule?

Answer 9

starting materials: CO2, glutamine, aspartate (where most atoms come from) and PRPP
the goal is to make UMP

Question 10

pyrimidine pathway cube

Answer 10

Question 11

what are the bases that need to be kept strictly on their corresponding sugars?

Answer 11

uracil only on ribose
thymine only on deoxyribose
put U into DNA -> mutations bc of repair or synthesis

Question 12

how is dUTP prevented from being put into DNA?

Answer 12

specific UTPase hydrolyzes dUTP to decrease its concentration and reduce its incorporation

Question 13

where do the atoms on pyrimidines come from?

Answer 13

Question 14

what is made first when making pyrimidine nucleotides?

Answer 14

pyrimidine ring is assembled separately, then the PRPP is added at the end to make the nucleotide
(it is the other way around for purines)
charged intermediates have low membrane permeability

Question 15

what are the steps in the pyrimidine de novo pathway? (make UMP)

Answer 15

aspartate transcarbamoylase is a well studied example of allosteric regulation
activated by ATP, inhibited by CTP

Question 16

continuation of the pathway

Answer 16

Question 17

graphic of how the enzymes are arranged in animals

Answer 17

Question 18

how is CTP made?

Answer 18

made at the triphosphate level from UTP, the N is from glutamine

Question 19

how is the ketone on UTP turned into the amine on CTP?

Answer 19

Question 20

by which enzymes does the interconversion of mono di and triphosphates happen?

Answer 20

base specific nucleotide kinases
non specific nucleoside diphosphate kinase

Question 21

at which level are deoxy made?

Answer 21

at the NDP level
the enzyme is ribonucleotide reductase

Question 22

what is pyrimidine synthesis inhibited and activated by?

Answer 22

inhibition by pyrimidine nucleoside triphosphates (product inhibition)
activated by ATP and PRPP
regulation is different in bacteria and animals
bacteria: step 2
animals: step 1