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Flashcards in Metabolism 11 Deck (24):
1

What are the sources of the atoms in the purine ring structure?

The sources of the atoms in the purine ring structure are three amino acids, CO2,
and a tetrahydrofolate (FH4) derivative.

2

What is the precursor for purine biosynthesis? How is this precursor produced?

The precursor for purine biosynthesis is ribose-5-phosphate, produced by the
pentose-phosphate pathway.

3

How is the purine built?

What is the first formal step?

Second step? Enzyme?

When is the nucleotide IMP formed? What is its significance?

When is the committed and major regulation step?

The purine ring is built up atom by atom on top of the ribose.

The first formal step (step 1 below) is the activation of ribose-5-phosphate to give
PRPP, with the pyrophosphate coming from ATP. (PRPP: 5-phospho-alpha-Dribose-
1-pyrophosphoric acid)

The second step is the formation of 5-phosphoribosylamine, with the amino group
coming from the side-chain of glutamine. This is the committed step and the
major regulated step. Enzyme: glutamine PRPP amidotransferase.

The nucleotide IMP is formed (step 11), and becomes the precursor for both AMP
and GMP.

4

Describe purine biosynthetic pathway.

p 5/6

5

Draw a summary and regulation of purine biosynthesis.

p 7

6

What happens to AMP and GMP that are generated in purine biosynthesis ?

How does the concentration of its products compare to concentrations of monophosphates, nucleosides or free bases?

AMP and GMP are phosphorylated to give nucleoside 5-diphosphates (ADP, GDP) and nucleoside 5-triphosphates (ATP, GTP), which are present in higher concentrations than monophosphates, nucleosides, or free bases.

7

How might an antitumor agent work? Why?

Tumors are very dependent on the purine biosynthetic pathway to produce purine nucleotides for DNA and RNA synthesis. In contrast, most normal tissues are less dependent on de novo synthesis and instead recycle existing purine bases (see salvage pathway below). A drug that inhibits purine biosynthesis would be more toxic to tumor cells than to most normal cells.

8

Tumors are very dependent on the purine biosynthetic pathway to produce purine nucleotides for DNA and RNA synthesis. In contrast, most normal tissues are less dependent on de novo synthesis and instead recycle existing purine bases (see salvage pathway below). A drug that inhibits purine biosynthesis would be more toxic to tumor cells than to most normal cells.

Describe the mechanism of antitumor agent, 6-mercaptopurine.

6-Mercaptopurine is converted to a nucleotide (by the salvage pathway). The nucleotide (monophosphate) then inhibits the enzymes in the purine biosynthetic pathway that catalyze steps 2, 12a, 12b, and 13a. It is used to treat acute lymphocytic leukemia (ALL) and other cancers.

p 7

9

What are the sources of the atoms in the pyrimidine ring structure?

The sources of the atoms in the pyrimidine ring structure are glutamine, aspartate,
and CO2:

10

Describe pyrimidine nucleotide biosynthesis.

What is the regulated step? What catalyzes it?

What happens first?

What is the role of UMP?

The regulated step (1) is catalyzed by carbamoyl phosphate synthetase II.

The pyrimidine ring structure is formed first and then attached to a ribose-5-
phosphate. PRPP is utilized as the activated form of ribose-5-phosphate.

UMP is the precursor for synthesis of the other pyrimidine nucleotides.

11

What is Leflunomide? What condition does it treat? How?

Leflunomide, an immunosuppressive drug used to treat rheumatoid arthritis,
blocks pyrimidine biosynthesis by inhibiting dihydroorotate dehydrogenase.

12

Describe the synthesis of deoxythymidylate (dTMP) and mechanism of action of the antitumor drugs 5-fluorouracil and methotrexate:

From what is dTMP synthesized?

What happens to 5-fluorouracil?

Describe the dihydrofolate produced in this reaction. What happens to it?

dTMP is synthesized from dUMP and N5,N10-methylene tetrahydrofolate by thymidylate synthase.

5-Fluorouracil is converted to f-dUMP which then acts as a pseudosubstrate and covalently binds to, and inhibits, thymidylate synthase.

The dihydrofolate produced in this reaction is useless and actually toxic and must be converted back to tetrahydrofolate by dihydrofolate reductase in order to be used for purine nucleotide and deoxythymidylate synthesis.

13

What does methotrexate do?

Methotrexate is a potent competitive inhibitor of dihydrofolate reductase and increases dihydrofolate and causes an intracellular tetrahydrofolate deficiency.

14

Describe the formation of deoxyribose.

What enzyme is used as a cofactor?

1. Ribonucleotide reductase acts on ribonucleoside 5-diphosphates (ADP, GDP,
CDP, or UDP) to form the corresponding deoxyribonucleoside 5-diphosphates
(dADP, dGDP, dCDP, or dUDP).

2. In this reaction, the 2-hydroxyl group of ribose is replaced with a hydrogen atom.
The enzyme uses thioredoxin as a cofactor.

p 10

15

Describe the cell cycle and nucleotide biosynthesis:

At what phase does DNA synthesis occur?

When in the cycle do the concentrations of deoxyribonucleotides increase?

When in the cycle do the enzymes involved in purine and pyrimidine biosynthesis increase?

1. DNA synthesis occurs during the S phase of the cell cycle.

2. The concentrations of the deoxyribonucleotides required for DNA synthesis are low during most of the cell cycle but increase greatly
during S phase.

3. Several of the enzymes involved in purine and pyrimidine biosynthesis
increase during late G1/early S phase. Among these are ribonucleotide reductase and thymidylate synthase. The amounts of these two enzymes correlate with tumor growth rate.

16

How is DNA and RNA in diet and mRNA cells degraded? To give what? Draw.

The DNA and RNA in the diet and mRNA in cells are degraded by a combination of nucleases, nucleotidases, and nucleoside phosphorylases (along with some side reactions that modify adenine and cytosine) to give free pyrimidine bases (uracil and thymine) and purine bases (guanine and hypoxanthine)

p 11

17

What happens to most of free purine bases produced in cells from degradation of mRNA?

Describe this process.

Most of the free purine bases (approx 90%) produced in cells from degradation of mRNA are reutilized rather than degraded:

“Salvage Pathway”:
Enzyme: Hypoxanthine-guanine phosphoribosyltransferase (HGPRTase)

hypoxanthine + PRPP IMP + PPi
guanine + PRPP GMP + PPi

18

What will result from a genetic deficiency in HGPRTase (Lesch-Nyhan syndrome)

A genetic deficiency in HGPRTase (results in Lesch-Nyhan syndrome which is characterized by neurological problems including spasticity, mental retardation, and a compulsive self-mutilation, affecting only males. Lesch-Nyhan patients also have gout.

19

Of the purine bases that are produced by catabolism of cellular mRNA, what percent is metabolized to uric acid?

Of the purine bases that are produced by catabolism of cellular mRNA, about 10% is metabolized to uric acid.

20

What is the final product of purine degradation? What happens? Describe this process.

Uric acid is the final product of purine degradation and is excreted. Xanthine oxidase catalyzes both the conversion of hypoxanthine to xanthine and xanthine to uric acid:

p 12

21

What two forms does uric acid exist in?

What is the predominant form at pH of blood?

Both forms have low solubility but which form is less soluble?

Uric acid exists in two tautomeric forms (enol and keto forms), one of which can be deprotonated to form urate (pKa = 5.8).

At the pH of blood, urate is the predominant form, while at the pH of urine, substantial amounts of uric acid exist. (Both forms have low solubility, but uric acid is less soluble than urate.)

22

What does excessive production or underexcretion of uric acid lead to?

Excessive production or underexcretion of uric acid can lead to gout:

Hyperuricemia (elevated urate in blood) can lead to deposition of sodium urate crystals in joints. The resulting tophi can elicit painful acute inflammatory responses.

Hyperuricosuria (elevated uric acid in urine) can lead to deposition of uric acid stones in the kidney.

23

Describe the drug allopurinol. What does it treat? How?

One important drug for the treatment of gout is allopurinol, an inhibitor of xanthine oxidase. Allopurinol and its metabolite oxypurinol block the formation of xanthine from hypoxanthine and uric acid from xanthine (see above). This reduces urate and uric acid in blood and urine, resp., and shrinks tophi and renal uric acid stones.

24

What can folate deficiency lead to?

megaloblastic anemia
See slides 1-5