Suppression of Δlon phenotypes in Escherichia coli by N‐terminal DnaK peptides

Abstract

Δlon mutant of Escherichia coli becomes hypersensitive to DNA damaging agents and over-produce capsule due to stabilization of the Lon substrates, namely, SulA and RcsA, respectively. These phenotypes were earlier found to be suppressed in Δlon ssrA::cat/pUC4 K and Δlon faa (DnaJ, G232D) strains, called as "Alp" strains. We observed that a plasmid carrying an E. coli chromosomal fragment harboring few genes, a heat shock gene htpY and a portion of dnaK capable of encoding truncated Nterminal ATPase domain (244 aa) could suppress lon mutant phenotypes. Deletion of htpY did not affect the efficiency of suppression. Clones expressing DnaK′ (244 aa) peptide alone could suppress both Δlon phenotypes in copy number dependent manner. Inactivation of clpQ did not affect the MMS R phenotype of Δlon strain carrying dnaK′ clones indicating that ClpYQ protease does not degrade SulA. We hypothesize that the high levels of defective DnaK′-DnaJ chaperone complex formed in these strains might lead to aggregation of SulA and RcsA and, thereby the suppression of Δlon phenotypes. Systematic deletion analysis of dnaK′ revealed that, ∼220 aa N-terminal DnaK peptide is required for suppression of cps-lac overexpression and ∼169 aa peptide is enough for the suppression of MMS S phenotype of Δlon mutant. K E Y W O R D S Δlon mutant, ATPase domain, DnaK, DnaJ, Escherichia coli, SulA Abbreviations: Alp, alternate lon protease; faa, function affecting alp; MMS, methyl methane sulphonate. 302 |

phz1 contributes much more to phenazine‐1‐carboxylic acid biosynthesis than phz2 in Pseudomonas aeruginosa rpoS mutant

“…All bacterial strains were inoculated in flasks containing 150 ml LB or GA broth at 37°C with shaking at 180 rpm. Samples were harvested once every 12 h. β‐Galactosidase‐specific activities were quantitatively determined according to the method of Miller by using sodium dodecyl sulfate and chloroform‐treated cells in appropriate amounts .…”