Sequence-selective alkylation and cross-linking induced by mitomycin C upon activation by DT-diaphorase.
Aerobic reduction of MMC by DTD, an obligate two-electron reductase, or chemical reduction by sodium borohydride results predominantly in monoalkylation of DNA at the guanine N7 position within 5'-GG-3' and 5'-GTC-3' sequences. The level of guanine N7 alkylation after DTD reduction increased as the pH was decreased from 7.8 and was optimal at pH 6.6. A similar profile of alkylation was obtained when the major metabolite of DTD-mediated MMC metabolism, 2,7-diaminomitosene, was further reduced by DTD. The sequence preference for DNA interstrand cross-linking (ISC) was also determined using singly end-labeled oligonucleotide duplexes. Reduction of MMC by DTD induced DNA cross-links which were resistant to piperidine cleavage. Exposure of cross-linked DNA to dimethyl sulfate or formic acid and subsequent piperidine cleavage displayed a discontinuity in band pattern which suggested a 5'-CG-3' preference for DNA ISC. Major groove alkylation is proposed to occur via generation, and subsequent metabolism by DTD, of 2,7-diaminomitosene. Cross-linking of DNA, at 5'-CG-3' sequences, is proposed to require the formation of either the protonated leucomitomycin C or the leucoaziridinomitosene during DTD-mediated metabolism of MMC.
Prakash
A S
AS
School of Pharmacy, University of Southern California, Los Angeles 90033.
Beall
H
H
Ross
D
D
Gibson
N W
NW
D000332:Aerobiosis; D000478:Alkylation; D001483:Base Sequence; D003432:Cross-Linking Reagents / Q000378:metabolism / Q000494:pharmacology*; D004247:DNA / Q000378:metabolism; D016685:Mitomycin / Q000378:metabolism* / Q000494:pharmacology*; D008969:Molecular Sequence Data; D016660:NAD(P)H Dehydrogenase (Quinone) / Q000378:metabolism*; D010957:Plasmids; D015183:Restriction Mapping
D003432:Cross-Linking Reagents; D016685:Mitomycin; D004247:DNA; D016660:NAD(P)H Dehydrogenase (Quinone)