Hydrogen peroxide and coffee induce G:C→T:A transversions in the lacI gene of catalase-defective Escherichia coli

Abstract

The mutagenicity of hydrogen peroxide (H₂O₂) was compared with that of coffee, a complex mixture which generates H₂O₂. An Escherichia coli strain defective in catalase activity (katG katE double mutant) and carrying a single copy mucAB (pRW144) plasmid was constructed to enhance the mutagenic response to oxidants. The ability of the mucAB genes to influence the type, frequency and distribution of H₂O₂-induced mutations was also investigated in isogenic bacteria lacking pRW144. Induced mutational spectra were characterized and compared with that of spontaneous mutagenesis. A total of 444 independent forward mutations affecting the first 210 bp of the lacI gene were identified by DNA sequence analysis. The spontaneous mutation spectrum showed no bias (P = 0.52) for substitutions at G:C base pairs. In contrast, in the H₂O₂-induced spectrum substitutions occurred preferentially at G:C base pairs (P < 0.0001) with a preponderance of G:C→T:A transversions (43.4% of H₂O₂-induced mutants versus 17.3% of spontaneous mutants). These data support the view that 7,8-dihydro-8-oxoguanine is the main premutagenic lesion induced by H₂O₂ and that catalase-defective bacteria have elevated levels of 8-oxoguanine in chromosome DNA after H₂O₂ exposure. Coffee produced a similar distribution of mutational events as H₂O₂ (P > 0.05), suggesting that this compound may be the main cause of the coffee-induced mutagenesis. The presence of plasmid pRW144 did not affect the frequency of H₂O₂-induced G:C→T:A transversions, but caused an increase in A:T→T:A transversions and a decrease in –1 base frameshifts. Although the frequencies of G:C→T:A transversions were similar in all three induced spectra (H₂O₂ and coffee ± pRW144), differences were observed in location of mutations throughout the target gene.