DNA mismatch-repair in Escherichia coli counteracting the hydrolytic deamination of 5-methyl-cytosine residues

Abstract

Derivatives of phage M13 were constructed and used for the in vitro preparation of heteroduplex DNA molecules containing base/base mismatches that mimick DNA lesions caused by hydrolytic deamination of 5-meC residues in Escherichia coli DNA (i.e. they carry a T/G mismatch in the special sequence context provided by the recognition site -CCA/TGG-of the Dcm-methyltransferase). Upon introduction of these heteroduplex DNAs into CaCl2-treated E. coli cells, the mismatches are efficiently repaired with high bias in favour of the DNA strand containing the mismatched guanine residue. This special DNA mismatch-repair operates on fully dam-methylated DNA and is independent of gene mutH. It thus fulfills the salient requirements of a repair pathway responsible for counteracting the spontaneous hydrolytic deamination of 5-meC in vivo. The repair efficiency is boosted by a 5-methyl group present on the cytosine residue at the next-nearest position to the 5' side of the mismatched guanine. The repair is severely impaired in host strains carrying a mutation in any of the three loci dcm, mutL and mutS.