Alteration of Ultraviolet Sensitivity of Thymidine Kinase by Allosteric Regulators, Normal Substrates, and a Photo-Affinity Label, 5-Iodo-2'-deoxyuridine, a Metabolic Analog of Thymidine

摘要: Abstract Ultraviolet irradiation of thymidine kinase with light primarily 253.7 nm wave length results in a loss catalytic activity at rate that follows first order inactivation kinetics. does not alter the sigmoidal nature ATP saturating curve, sensitivity enzyme to allosteric regulators (dCDP or dTTP), Km for and 5-iodo-2'-deoxyuridine. Catalytic is restored by postirradiation incubation substrate. The ultraviolet altered presence molecules interact this affect its activity. Regulatory deoxynucleotides (dTTP, dCDP) normal substrates (ATP, thymidine) reduce inactivation. photoreactive substrate 5-iodo-2'-deoxyuridine enhances but activities irradiation. effect concentration on extent protection closely parallels their formation thymidylate. enhancement observed hyperbolic function substrate, similar saturation curve displaced higher concentrations factor 2 3. can be prevented thymidine. Furthermore, protective may predictable fashion appropriate regulator; i.e. both effects are reduced dTTP which decreases enzymic affinity increased dCDP increases It postulated photolysis deoxyuridine-free radical when formed halogenated enzyme-substrate complex causes modification one more amino acids active site vital