Comparison of mutagenesis by O6-methyl- and O6-ethylguanine and O4-methylthymine in Escherichia coli using double-stranded and gapped plasmids

Carcinogenesis
Gary T PaulyRobert C Moschel

Abstract

To compare mutagenesis by O6-methylguanine (m6G), O4-methylthymine (m4T) and O6-ethylguanine (e6G), and assess their genotoxicity in Escherichia coli, double-stranded and gapped plasmids were constructed containing a single m6G, e6G or m4T in the initiation codon (ATG) of a lacZ' gene. Modified base induced mutations were scored by the loss of lacZ' activity on X-gal-containing media resulting in formation of white or sectored (mutant) rather than blue (non-mutant) colonies. Genotoxicity experiments with gapped plasmids containing the modified bases indicated that m4T produced a greater number of bacterial colonies than m6G or e6G. m4T was more mutagenic (45% mutant colonies) than m6G (6%) or e6G (11%) in repair competent (w.t.) E. coli when incorporated in double-stranded plasmids. In gapped plasmids, m4T produced 99% mutant colonies (as was observed previously for e6G) in both w.t. E. coli or E. coli deficient in both O6-alkylguanine-DNA alkyltransferases as well as methylation-directed mismatch repair (ada(-)-ogt(-)-mutS[-]). m6G in gapped plasmids produced 62% mutant colonies in w.t. E. coli, but this percentage increased to 94% in the ada(-)-ogt(-)-mutS(-) strain. In double-stranded plasmids both m4T and m6G produced very ...Continue Reading

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