Novel mutations of Escherichia coli that produce recombinogenic lesions in DNA : V. Recombinogenic plasmids from arl mutants of Escherichia coli are unusually sensitive to nuclease S 1 and partially deficient in cytosine methylation at C-C-(A/T)-G-G sequences

Abstract

Two novel phenotypes previously associated with arl mutations of Escherichia coli, increased frequencies of genetic recombination and unusual sensitivity of DNA to the single-strand-specific nuclease S 1, have been defined most completely by the properties of λ bacteriophages grown on arl bacteria (Arl − phages). We now find that plasmids maintained in arl mutants (Arl − plasmids) exhibit elevated recombination frequencies, unusual sensitivity to nuclease S 1 (in a limited number of regions) and a new Arl phenotype, partially deficient methylation of the inner cytosine at C-C-(A/T)-G-G sequences. A variety of Arl − plasmids (all pBR322 derivatives) show elevated recombination (4 to 10-fold) by three different assays (frequencies of homomultimers and of heteromultimers, efficiency of intramolecular recombination). Plasmids from arl bacteria (after conversion to linear form) are nicked by nuclease S 1 about 0.7 times per duplex; Arl + plasmids are nuclease S 1-resistant. Restriction endonuclease EcoRII (recognition sequence, C-C-(A/T)-G-G) cuts Arl − plasmid DNA more readily than Arl + DNA, but Arl − plasmids are still more EcoRII-resistant than Dcm − plasmids (from E. coli dcm mutants, which lack the chromosomal cytosine methylase; recognition sequence, also C-C-(A/T)-G-G). By chromatographic analyses, Arl − plasmid DNA contains less 5-methylcytosine than Arl + (0.07% versus 0.15%). although the 6-methyladenine content is the same (0.5mol%).