Repair of methyl methane sulfonate-damaged phage by Haemophilus influenzae

Summary

Seven mutants of Haemophilus influenzae strain Rd (mmsA ^-) have been isolated that are more sensitive to methyl methane sulfonate (mms) than recombination-deficient (recA ^-) mutants. The mutations cotransformed about 25% with the strA locus while the five studied clustered tightly; they are all probably allelic. The mutants are not sensitive to ultraviolet radiation, X-rays, or nitrous acid. Mms-damaged phage HP1 plated very inefficiently on these mutants, indicating that they lack the first step in the excision repair of the lesion N3-methyladenine (m³A). Incubation of damaged phage at 30°C in the absence of mms resulted in a steady decline of viability when the phage were plated on the wild mmsA ⁺ host but an initial steep rise was seen when it was plated on an mmsA ^- mutant. The rise is explained by the assumption that m³A lesions hydrolyzed off the DNA giving rise to repairable apurinic sites by both the mmsA ⁺ and mmsA ^- hosts. No decline in viability was observed when hydroxylamine was present in the medium. This compound is known to prevent or slow down β-elimination. The delayed decline in viability is therefore explained by assuming that apurinic sites give rise to β-elimination-induced single strand breaks in the phage DNA that cannot be repaired by either host. Marker rescue experiments indicated that these breaks did not interrupt injection of phage DNA.