Characterization of Organic Hydroperoxide Resistance Gene VPA1681 against Organic Peroxides in the Presence of Alkylhydroperoxide Reductase Genes in Vibrio parahaemolyticus

The marine foodborne enteropathogen Vibrio parahaemolyticus contains the chief organic peroxide reductases AphC1-AhpC2 and a putative organic hydroperoxide resistance enzyme (Ohr, VPA1681) against different peroxides. This study investigated the function of the Ohr under the presence of AhpC1-AhpC2 in this pathogen by gene mutation. Experimental results demonstrated that the ohr gene product was a weak scavenger of H 2 O 2 only in the mutant strains that lacked the peroxide sensor/regulator oxyR and ahpC1 - ahpC2 genes. The Ohr of V. parahaemolyticus was highly effective in scavenging organic peroxide, as demonstrated by assaying the defective changes in the △ ohr mutant strain and determining the detoxifying activity of the purified recombinant V. parahaemolyticus Ohr vp protein in the reduced form. The Ohr and AhpC1-AhpC2 exhibited similar functions against organic peroxides; however, only the △ ahpC1△ahpC2 mutant strain showed a significant increase in susceptibility to several disinfectants, organic acids, and antibiotics compared to the wild-type strain. The transcription of the ohr gene depended on exogenous cumene hydroperoxide (cumene) stress and was markedly enhanced in the △ ohrR (VPA1682) mutant strains. This study revealed the organic hydroperoxide reductase activity of the Ohr in V. parahaemolyticus , and its role probably depends on the sophisticated regulation by OhrR. IMPORTANCE Vibrio parahaemolyticus is the most prevalent foodborne pathogen in Taiwan and some other coastal Asian countries, and its antioxidative activity contributes to the tolerance of this bacterium to different environmental stresses. This study reports on the function of the organic hydroperoxide resistance gene ( ohr , VPA1681) and its gene regulator ohrR (VPA1682) in this pathogen. The strain with ohr gene was effective in protection against organic peroxide, and the recombinant Ohr vp was active in its reduced form. The function of Ohr was significant mostly in strains in which the function o f AhpC1-AhpC2 was limited. The ohrR repressor of the ohr gene was effective at low concentrations of organic peroxide. Other common Vibrio species contain homologous ohr , ohrR , ahpC1, and ahpC2 genes, which are phylogenetically close to those of V. parahaemolyticus may probably share similar functions to those revealed in this study.

Ohr and OhrR Are Critical for Organic Peroxide Resistance and Symbiosis in Azorhizobium caulinodans ORS571

Azorhizobium caulinodans is a symbiotic nitrogen-fixing bacterium that forms both root and stem nodules on Sesbania rostrata. During nodule formation, bacteria have to withstand organic peroxides that are produced by plant. Previous studies have elaborated on resistance to these oxygen radicals in several bacteria; however, to the best of our knowledge, none have investigated this process in A. caulinodans. In this study, we identified and characterised the organic hydroperoxide resistance gene ohr (AZC_2977) and its regulator ohrR (AZC_3555) in A. caulinodans ORS571. Hypersensitivity to organic hydroperoxide was observed in an ohr mutant. While using a lacZ-based reporter system, we revealed that OhrR repressed the expression of ohr. Moreover, electrophoretic mobility shift assays demonstrated that OhrR regulated ohr by direct binding to its promoter region. We showed that this binding was prevented by OhrR oxidation under aerobic conditions, which promoted OhrR dimerization and the activation of ohr. Furthermore, we showed that one of the two conserved cysteine residues in OhrR, Cys11, was critical for the sensitivity to organic hydroperoxides. Plant assays revealed that the inactivation of Ohr decreased the number of stem nodules and nitrogenase activity. Our data demonstrated that Ohr and OhrR are required for protecting A. caulinodans from organic hydroperoxide stress and play an important role in the interaction of the bacterium with plants. The results that were obtained in our study suggested that a thiol-based switch in A. caulinodans might sense host organic peroxide signals and enhance symbiosis.

Photocatalysed decolouration of indigo in solution via in situ generation of an organic hydroperoxide

Indigo, a famous and well-loved dye, is highly resistant towards photobleaching but can be broken down to the colourless isatin under photocatalytic conditions.

Gene Regulation by Redox-Sensitive Burkholderia thailandensis OhrR and Its Role in Bacterial Killing of Caenorhabditis elegans

ABSTRACT Fatty acid hydroperoxides are involved in host-pathogen interactions. In both plants and mammals, polyunsaturated fatty acids are liberated during infection and enzymatically oxidized to the corresponding toxic hydroperoxides during the defensive oxidative burst that is designed to thwart the infection. The bacterial transcription factor OhrR (organic hydroperoxide reductase regulator) is oxidized by organic hydroperoxides, as a result of which the ohr gene encoding organic hydroperoxide reductase is induced. This enzyme converts the hydroperoxides to less toxic alcohols. We show here that OhrR from Burkholderia thailandensis represses expression of ohr. Gene expression is induced by cumene hydroperoxide and to a lesser extent by inorganic oxidants; however, Ohr contributes to degradation only of the organic hydroperoxide. B. thailandensis OhrR, which binds specific sites in both ohr and ohrR promoters, as evidenced by DNase I footprinting, belongs to the 2-Cys subfamily of OhrR proteins, and its oxidation leads to reversible disulfide bond formation between conserved N- and C-terminal cysteines in separate monomers. Oxidation of the N-terminal Cys is sufficient for attenuation of DNA binding in vitro, with complete restoration of DNA binding occurring on addition of a reducing agent. Surprisingly, both overexpression of ohr and deletion of ohr results in enhanced survival on exposure to organic hydroperoxide in vitro. While Δohr cells are more virulent in a Caenorhabditis elegans model of infection, ΔohrR cells are less so. Taken together, our data suggest that B. thailandensis OhrR has several unconventional features and that both OhrR and organic hydroperoxides may contribute to virulence.