Thymidylate synthase-catalyzed, tetrahydrofolate-dependent self-inactivation by 5-FdUMP.

Thymidylate synthase-catalyzed, tetrahydrofolate-dependent self-inactivation by 5-FdUMP.
Arch Biochem Biophys. 2019 Sep 11;:108106
Authors: Sobich J, Prokopowicz M, Maj P, Wilk P, Zieliński Z, Frączyk T, Rode W
Abstract
In view of previous crystallographic studies, N4-hydroxy-dCMP, a slow-binding thymidylate synthase inhibitor apparently caused "uncoupling" of the two thymidylate synthase-catalyzed reactions, including the N5,10-methylenetetrahydrofolate one-carbon group transfer and reduction, suggesting the enzyme's capacity to use tetrahydrofolate as a cofactor reducing the pyrimidine ring C(5) in the absence of the 5-methylene group. Testing the latter interpretation, a possibility was examined of a TS-catalyzed covalent self-modification/self-inactivation with certain pyrimidine deoxynucleotides, including 5-fluoro-dUMP and N4-hydroxy-dCMP, that would be promoted by tetrahydrofolate and accompanied with its parallel oxidation to dihydrofolate. Electrophoretic analysis showed mouse recombinant TS protein to form, in the presence of tetrahydrofolate, a covalently bound, electrophoretically separable 5-fluoro-dUMP-thymidylate synthase complex, similar to that produced in the presence of N5,10-methylenetetra-hydrofolate. Further studies of the mouse enzyme binding with 5-fluoro-dUMP/N4-hydroxy-dCMP by TCA precipitation of the complex on filter paper showed it to be tetrahydrofolate-promoted, as well as to depend on both time in th...

Source: Archives of Biochemistry and Biophysics - September 10, 2019 Category: Biochemistry Authors: Sobich J, Prokopowicz M, Maj P, Wilk P, Zieliński Z, Frączyk T, Rode W Tags: Arch Biochem Biophys Source Type: research