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)


The de novo synthesis of thymidylate from deoxyuridylate proceeds by the following sequence of reactions:
$$ 5,10-Methylenetetrahydrofolate + deoxyuridylate\to dihydrofolate + thymidylate $$
$$ Dihydrofolate + TPNH+{{H}^{+}}\to tetrahydrofolate + TP{{N}^{+}} $$
$$ Tetrahydrofolate + "- C{H_2} -" \to 5,10 - methylenetetrahydrofolate $$
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.


Affinity Chromatography Snake Venom Column Material Thymidylate Synthetase Phosphate Derivative 
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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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