Lecture 33 Summary

The synthesis of DNA requires the prior synthesis of deoxyribonucleotides. This is accomplished by the action of ribonucleotide reductase. Reducing power from NADPH is transferred to ribonucleotide reductase via two separate carrier systems in E. coli. One involves thioredoxin, the other uses glutathione.

Uridylate must also be methylated at C5 to yield thymidylate for DNA synthesis. The methylation is carried out by the enzyme thymidylate synthase. N⁵-N¹⁰-methylenetetrahydrofolate (N⁵-N¹⁰-THF), a folate cofactor is the methyl donor for this reaction. The cofactor must be regenerated via a cycle completed by a reductase and the transfer of a methylene group from serine. Both thymidylate synthase and dihydrofolate reductase have been the targets of anticancer drugs.

Ribonucleotide reductase [EC 1.17.4.1] acts on NTP substrates. Structure and function of ribonucleotide reductase. Ribonucleotide reductases act via a radical mechanism. Production of thymidylate from dUMP. Anticancer drugs and the dUMP/methylfolate cycle

Thymidylate synthase [TS, EC 2.1.1.45] is the enzyme that catalyzes the addition of a methyl group to the deoxyribonucleotide uridylate (dUMP) to form thymidylate (dTMP). The one-carbon unit is donated by N⁵,N¹⁰-methylenetetrahydrofolate, which is also oxidized in the process to dihydrofolate. Since the production of dTMP is required for the synthesis of DNA, and the rapidly proliferating cells of cancerous tissue are heavily dependent on DNA synthesis, TS has been a target of inhibitory drugs that act as chemotherapeutic agents. One such agent is 5-fluorodeoxyuridylate (FdUMP), a mechanism-based or suicide inhibitor of TS.

Study questions

• Which anticancer drug would you expect to have greater side effects, fluorouracil or methotrexate? Explain.

Page updated 12-27-06

References

1. Berg, Tymoczko, and Stryer. Biochemistry (BTS): 6th edition (2007, Freeman) Ch.25, pp.718-728.