Kinetic and Photochemical Studies of 3-N-Methyl-5-iodo-2′-deoxyuridine

The effects of 5-iodo-2'-deoxyuridine, 5-bromo-2'-deoxyuridine, 5-chloro-2'-deoxyuridine, 5-fluoro-2'-deoxyuridine, and 3-N-methyl-5-iodo-2'-deoxyuridine, a new analog of thymidine, on the ultraviolet (UV) inactivation of thymidine kinase have been investigated. Of these compounds only 5-iodo-2'-deoxyuridine and 3-N-methyl-5-iodo-2'-deoxyuridine enhance the rate of inactivation of the enzyme by UV light. The other halogenated analogs neither protect nor sensitize thymidine kinase to UV inactivation. Whereas the inactivation of thymidine kinase by UV light in the presence of 5-iodo-2'-deoxyuridine can be prevented by the substrate, thymidine (Cysyk, R., and Prusoff, W. H. (1969) Fed. Proc., 28, 473; Cysyk, R. (1970) Ph.D. thesis, Yale University), that produced by 3-N-methyl-5-iodo-2'-deoxyuridine is not prevented by thymidine alone but requires the presence of both substrates, thymidine and MgATP. Kinetic studies with thymidine kinase show 5-iodo-2'-deoxyuridine to be a competitive inhibitor with respect to thymidine; however, 3-N-methyl-5-iodo-2'-deoxyuridine shows uncompetitive inhibition with thymidine and competitive inhibition with MgATP. The allosteric regulators, dCDP, an activator, and dTTP, an inhibitor, protect the enzyme against UV irradiation and decrease the rate of inactivation caused by 3-N-methyl-5-iodo-2'-deoxyuridine. The primary photostable compound formed during photolysis of 3-N-methyl-5-iodo-2'-deoxyuridine appears to be 3-N-methyl deoxyuridine, which is analogous to the formation of deoxyuridine during photolysis of 5-iodo-2'-deoxyuridine (Cysyk, R. (1970) Ph.D. thesis, Yale University).