Characterization of a dam mutant of Haemophilus influenzae Rd

Abstract

The gene encoding Dam methyltransferase ofHaemophilus influenzaewas mutagenized by the insertion of a chloramphenicol-resistance cassette into the middle of the Dam coding sequence. This mutant construct was introduced into theH. influenzaechromosome by transformation and selection for CamRtransformants. The authors have shown that several phenotypic properties, resistance to antibiotics, dyes and detergent as well as efficiency of transformation, depend on the Dam methylation state of the DNA. Although the major role of the methyl-directed mismatch repair (MMR) system is to repair postreplicative errors, it seems that inH. influenzaeits effect is more apparent in repairing DNA damage caused by oxidative compounds. In thedammutant treated with hydrogen peroxide, MMR is not targeted to newly replicated DNA strands and therefore mismatches are converted into single- and double-strand DNA breaks. This is shown by the increased peroxide sensitivity of thedammutant and the finding that the sensitivity can be suppressed by amutHmutation inactivating MMR. In thedammutant treated with nitrofurazone the resulting damage is not converted into DNA breaks but the high sensitivity is also suppressed by amutHmutation.