Abstract
Enzyme kinetic parameters of the bioactivation of thiourea-containing compounds by human flavin-containing monooxygenase enzymes (FMOs) FMO1 and FMO3 were investigated. A microtitre-based adaptation of methodology described for the thiourea-dependent oxidation of thiocholine was used to determine the turnover of thiourea-containing compounds by human FMO1 and FMO3. The results show that major differences in enzyme kinetic parameters for N-substituted N′-(4-imidazole-ethyl)thiourea exist between human FMO3 and human FMO1. Whereas Km values of N-substituted N′-(4-imidazole-ethyl)thioureas for human FMO3 are all in the millimolar range, the Km values for human FMO1 range from the low micromolar to the low millimolar range. Furthermore, among a series of N-p-phenyl-substituted N′-(4-imidazole-ethyl)thioureas an interesting structure–activity relationship is evident with both FMO1 and FMO3. Where the Km decreases with increasing electron-withdrawing capacity of the p-substituent in the case of FMO1, the opposite phenomenon may be the case with FMO3. The kcat values of the compounds were all comparable for FMO1, averaging 3.03 ± 0.56 min−1, whereas more variation was found for FMO3 (3.71 ± 2.01 min−1). Enzyme kinetic parameters Km and kcat/Km of human FMO1 for N-substituted N′-(4-imidazole-ethyl)thioureas show a high degree of correlation with the results obtained in rat liver microsomes, in which rat FMO1 is the most abundant form, whereas those of human FMO3 do not.
Acknowledgement
Work was supported in part by NIH Grant GM43511 (A.E.R.).