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Affinity Chromatography of Thymidylate Synthetases Using 5-Fluoro-2′-Deoxyuridine 5′-Phosphate Derivatives of Sepharose

  • John M. Whiteley
  • Ivanka Jerkunica
  • Thomas Deits
Part of the Advances in Experimental Medicine and Biology book series (AEMB, volume 42)

Abstract

The de novo synthesis of thymidylate from deoxyuridylate proceeds by the following sequence of reactions:
$$ 5,10-Methylenetetrahydrofolate + deoxyuridylate\to dihydrofolate + thymidylate $$
(1)
$$ Dihydrofolate + TPNH+{{H}^{+}}\to tetrahydrofolate + TP{{N}^{+}} $$
(2)
$$ Tetrahydrofolate + "- C{H_2} -" \to 5,10 - methylenetetrahydrofolate $$
(3)
In the presence of thymidylate synthetase, 5,10-methylenetetrahydrofolate donates its labile one-carbon unit (the methylene group) to deoxyuridylate, and also provides the reducing power necessary to convert the methylene group to methyl leading to the formation of thymidylate and dihydrofolate, as is shown in equation (1). The dihydrofolate so formed is converted to tetrahydrofolate with the aid of a second enzyme, dihydrofolate reductase (equation (2)). The cycle is then completed (equation (3)) by tetrahydrofolate acquiring the one-carbon unit necessary to re-form the methylene derivative utilized in the first reaction. Because thymidylate is essential for the biosynthesis of DNA, these reactions play a leading role in cellular replication. This factor is probably responsible for the potent chemotherapeutic properties possessed by compounds such as amethopterin and 5-fluorouracil.

Keywords

Affinity Chromatography Snake Venom Column Material Thymidylate Synthetase Phosphate Derivative 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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References

  1. 1.
    Wahba, A.J., and Friedkin, M., J. Biol. Chem. 236, PC11 (1961).Google Scholar
  2. 2.
    Pastore, E.J., and Friedkin, M., J. Biol. Chem. 237, 3802 (1962).Google Scholar
  3. 3.
    Reyes, P., and Heidelberger, C., Mol. Pharmacol. 1, 14 (1965).Google Scholar
  4. 4.
    Blakley, R.L., J. Biol. Chem. 238, 2113 (1963).Google Scholar
  5. 5.
    Crusberg, T.C., and Kisliuk, R.L., Fed. Proc. 28, 473 (1969).Google Scholar
  6. 6.
    Dunlap, R.B., Harding, N.G.L., and Huennekens, F.M., Biochemistry 10, 88 (1971).CrossRefGoogle Scholar
  7. 7.
    Whiteley, J.M., Jackson, R.C., Mell, G.P., Drais, J.H., and Huennekens, F.M., Arch. Biochem. Biophys. 150, 15 (1972).CrossRefGoogle Scholar
  8. 8.
    Dann, J.G., Harding, N.G.L., Newbold, P.C.H., and Whiteley, J.M., Biochem. J. 127, 28P (1972).Google Scholar
  9. 9.
    Leary, R.P., and Kisliuk, R.L., Prep. Biochem. 1, 47 (1971).CrossRefGoogle Scholar
  10. 10.
    Cohen, S.S., Flaks, J.G., Barner, H.D., Loeb, M.R., and Lichenstein, J., Proc. Nat. Acad. Sci. (U.S.A.) 44, 1004 (1958).CrossRefGoogle Scholar
  11. 11.
    Axen, R., Porath, J., and Ernback, S., Nature (London) 214, 1302 (1967).CrossRefGoogle Scholar
  12. 12.
    Danenberg, P.V., Langenback, R.J., and Heidelberger, C., Biochem. Biophys. Res. Commun. 49, 1029 (1972).CrossRefGoogle Scholar
  13. 13.
    Whiteley, J.M., Jerkunika, I., and Deits, T., submitted to Biochemistry.Google Scholar
  14. 14.
    Tener, G.M., J. Amer. Chem. Soc. 83, 159 (1961).CrossRefGoogle Scholar
  15. 15.
    Cuatrecasas, P., Wilchek, M., and Anfinsen, C.B., Biochemistry 8, 2277 (1969).CrossRefGoogle Scholar
  16. 16.
    Chen, H.-C., Craig, L.C., and Stauer, E., Biochemistry 11, 3559 (1972).CrossRefGoogle Scholar
  17. 17.
    Galivan, J., Maley, G.F., and Maley, F., Fed. Proc. 32, 2116 (1973).Google Scholar

Copyright information

© Plenum Press, New York 1974

Authors and Affiliations

  • John M. Whiteley
    • 1
  • Ivanka Jerkunica
    • 1
  • Thomas Deits
    • 1
  1. 1.Scripps Clinic and Research FoundationLa JollaUSA

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