Co-induction of DNA relaxation and synthesis of DnaK and GroEL proteins in Escherichia coli by expression of LetD (CcdB) protein, an inhibitor of DNA gyrase encoded by the F factor.

Abstract

We examined the influence of overexpression of LetD (CcdB) protein, an inhibitor of DNA gyrase encoded by the F factor of Escherichia coli, on DNA supercoiling and induction of heat shock proteins. Cells were transformed with a plasmid carrying the structural gene for LetD protein under control of the tac promoter, and LetD protein was induced by adding isopropyl beta-D-thiogalactopyranoside (IPTG) to the culture medium. Analysis by agarose gel electrophoresis in the presence of chloroquine revealed relaxation of plasmid DNA in cells depending on the concentration of IPTG employed for induction. Protein pulse-labeling experiments with [35S]methionine and cysteine revealed that synthesis of DnaK and GroEL proteins was also induced by IPTG, and concentrations necessary for DNA relaxation and induction of the heat shock proteins were much the same. Expression of mutant LetD protein lacking two amino acid residues at the C-terminus induced neither DNA relaxation nor the synthesis of DnaK and GroEL proteins. Induction of wild-type LetD protein but not mutant LetD protein markedly enhanced synthesis of sigma32. We interpret these results to mean that DNA relaxation in cells caused by the expression of LetD protein induces heat shock proteins via increased synthesis of sigma32.