Mechanism of inhibition of erythrocyte glutathione reductase by mitomycin-C.
Mitomycin-C (MT-C) inhibits human erythrocyte glutathione reductase (EGR). We found that Km values of EGR for GSSG, NADPH, and FAD were 1.3 X 10(-4) mol/l, 2.3 X 10(-5) mol/l, and 3.9 X 10(-7) mol/l respectively, and that EGR was inhibited by MT-C non-competitively with respect to both the substrate GSSG (Ki = 4.8 X 10(-5) mol/l) and the cofactor NADPH (Ki = 2.2 X 10(-4) mol/l), and competitively with respect to FAD (Ki = 4.2 X 10(-5) mol/l). On the other hand, we demonstrated that FAD makes a complex with MT-C, the dissociation constant (K = 6.5 X 10(-5) mol/l) of which was obtained from fluorescence quenching of FAD with MT-C. It also became clear that spectra wf FAD at around 375 nm changes with increasing MT-C concentration. So, the data strongly suggest that the mechanism of inhibition of EGR by MT-C is mainly due to complex formation of the co-enzyme FAD with MT-C.
Okuda
J
J
Department of Clinical Biochemistry, Faculty of Pharmacy, Meijo University, Nagoya, Japan.
Hirai
Y
Y
Hayazaki
T
T
D004912:Erythrocytes / Q000201:enzymology*; D005980:Glutathione Reductase / Q000037:antagonists & inhibitors* / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D016685:Mitomycin; D008937:Mitomycins / Q000494:pharmacology*; D009249:NADP / Q000378:metabolism; D010084:Oxidation-Reduction
D008937:Mitomycins; D016685:Mitomycin; D009249:NADP; D005980:Glutathione Reductase