MUTAGENESIS IN ESCHERICHIA-COLI K-12-MUTANTS DEFECTIVE IN SUPEROXIDE-DISMUTASE OR CATALASE

Escherichia coli K-12 strains with diminished levels of superoxide dismutase (SOD) due to inactivation of the sodA, sodB or sodA sodB genes were constructed in order to quantify the role of O2-. in mutagenesis. Mutagenesis was monitored by selecting forward mutations to L-arabinose resistance (Ara(R)). No sodA sodB mutant inability to grow in aerobic minimal medium was found, in contrast to that previously reported for a different E.coli wild-type genetic background. The role of SOD for coping with the damaging effects of superoxide became evident after the increase in intracellular 02- . flux by growing cells under hyperoxygenation, but particularly by using redox cycling compounds such as plumbagin, paraquat and menadione. Bacteria completely devoid of SOD activity showed very high levels of Ara(R)-induced mutants at doses that were non-mutagenic for the SOD-proficient parental or the sodA or sodB single mutants. The mutagenicity of nifurtimox and quercetin were studied to further compare the responses of the SOD-deficient bacteria to those of their SOD-proficient counterparts. The relative importance of SOD and catalase for coping with the damaging effects of O2-. and H2O2 was quantified by comparing SOD-deficient bacteria with isogenic catalase-deficient cells (a katG katE double mutant). The mutagenicities of plumbagin and menadione were much higher in SOD-deficient than in catalase efficient bacteria, in agreement with the role of the O2-. radical in the so-called metal-catalyzed Haber-Weiss reaction. The relevance of catalase in protecting against the damaging effects of H2O2 was evident from the hypersensitivity of the katG katE double mutant to the mutagenic and lethal effects of this oxidizing agent. It is concluded that the Ara mutagenicity assay combined with depletion in specific antioxidative enzymes could be a tool in establishing the extent to which DNA damage by oxygen radicals is relevant to mutagenesis.