5-p-benzoquinonyl-2'-deoxyuridine 5'-phosphate: a possible mechanism-based inhibitor of thymidylate synthetase.

The title compound (1), designed as a suicide inhibitor of thymidylate synthetase, can be prepared by silver(II) oxide oxidative demethylation of the corresponding dimethoxyphenyl derivative. Compound 1 shows time-dependent inactivation of thymidylate synthetase (methotrexate-resistant Lactobacillus casei) and saturation kinetics, and the inactivation is responsive to substrate protection. The inactivation is not reversible on prolonged dialysis in attempts to remove the inhibitor. The rate constant for inactivation is 0.065 s-1; the dissociation constant (Ki) was estimated to be 2 microM. The kinetics of this inactivation are compared to inactivation caused by model thiol reagents that do not have affinity for the active site of thymidylate synthetase.