The Molecular Chaperone DnaJ is Involved in the Peroxide- induced Oxidative Stress Response in Escherichia coli

The chaperone protein DnaJ and sigma (σ) factor σ32 (often called RpoH) play important roles in the heat-shock response in Escherichia coli. Previous studies have demonstrated increased heat tolerance accompanied by increased RpoH concentrations in E. coli treated with a sub-minimal inhibitory concentrations (MIC) of kanamycin. DnaJ, along with other chaperone proteins, forms a complex with RpoH to facilitate its degradation in the absence of stressful stimuli. In this study, we explored the relative importance both proteins in the oxidative stress response in the presence and absence of a sub-MIC of kanamycin. Phagocytes of the human immune system often use reactive oxygen species upon contact with foreign bacteria to facilitate killing and prevent potential infection. Tolerance of E. coli strain C600, dnaJ mutant strain MF634, and ΔrpoH strain K165 to peroxide-induced oxidative stress was assessed by (i) monitoring cell growth in broth cultures containing H2O2 and by (ii) measuring zones of H2O2 growth inhibition in a solid phase diffusion disc assay. In broth cultures, growth of the wild-type strain was less negatively impacted by H2O2 compared to the dnaJ and ΔrpoH mutants. In the disc diffusion assay, zones of inhibition for WT were significantly smaller than those of the dnaJ mutant. The presence of a sub-MIC of kanamycin did not produce smaller zones of inhibition for either WT or mutant strains in the disc diffusion assay. Western blot detection analysis showed that in the absence of a sub-MIC of kanamycin, RpoH concentrations were higher in the dnaJ mutant compared to the WT strain. Results of this study indicate that DnaJ is an important mediator of the oxidative stress and that a lack of a functional copy of dnaJ is sufficient to cause impairment of the oxidative stress response in E. coli.