Metabolic activation of mitomycin C by NADPH-ferredoxin reductase in vitro.
Mammalian NADPH-ferredoxin reductase (EC 1.18.1.2) functions in the mitochondrial electron transport chain for cytochrome P-450-dependent steroid hydroxylation. Significant homology of three-dimensional structure exists in the surroundings of FAD between NADPH-ferredoxin reductase and NADH-cytochrome b5 reductase. The latter is involved in the bioreduction of mitomycin C (MC), a prototype antitumor agent. In this study, we assessed the capacity of NADPH-ferredoxin reductase to activate MC. Mitomycin C increased the NADPH oxidase activity of NADPH-ferredoxin reductase. In the absence of ferredoxin, the Km value of NADPH-ferredoxin reductase for MC was 73.5 +/- 2.3 microM. While in the presence of 500 nM ferredoxin, a Lineweaver-Burk plot exhibited a biphasic curve. NADPH-ferredoxin reductase-mediated reduction of MC resulted in the formation of an alkylated complex of 4-(p-nitrobenzyl) pyridine and an increase in plasmide DNA single-strand breaks under hypoxic conditions. With the addition of 500 nM ferredoxin, the amount of the alkylated complex of 4-(p-nitrobenzyl) pyridine and the plasmide DNA single-strand breaks increased by 40% and 37%, respectively. However, neither alkylated complex of 4-(p-nitrobenzyl) pyridine nor DNA strand breaks was observed in the presence of SOD and catalase under aerobic conditions. These findings demonstrate that NADPH-ferredoxin reductase is capable of catalyzing the bioactivation of mitomycin C under hypoxic conditions in vitro.
Department of Biochemistry, Kagawa Medical University, Kita-gun, Japan.
H B
HB
Jiang
M
M
Ichikawa
A
A
Furukawa
S
S
Tomita
T
T
Ohnishi
Y
Y
Ichikawa
D000478:Alkylation; D000818:Animals; D000903:Antibiotics, Antineoplastic / Q000378:metabolism*; D001711:Biotransformation / Q000502:physiology; D002417:Cattle; D004249:DNA Damage; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005287:Ferredoxin-NADP Reductase / Q000235:genetics / Q000378:metabolism*; D000860:Hypoxia / Q000473:pathology; D016685:Mitomycin / Q000378:metabolism*; D010084:Oxidation-Reduction
D000903:Antibiotics, Antineoplastic; D016685:Mitomycin; D005287:Ferredoxin-NADP Reductase