Molecular Insights into Hydrogen Peroxide Sensing Mechanism of the Metalloregulator MntR in Controlling Bacterial Resistance to Oxidative Stresses [Gene Regulation]

Manganese contributes to anti-oxidative stress particularly in catalase-devoid bacteria, and DtxR family metalloregulators, through sensing cellular Mn2+ content, regulate its homeostasis. Here, we show that metalloregulator MntR (So-MntR) functions dually as Mn2+ and H2O2 sensors in mediating H2O2 resistance by an oral streptococcus. H2O2 disrupted So-MntR's binding to Mn2+ transporter mntABC promoter, and induced disulfide-linked dimerization of the protein. Mass spectrometry identified Cys11-Cys156 and Cys11-Cys11 disulfide-linked peptides in H2O2 treated So-MntR. Site mutagenesis of Cys11 and Cys156, and particularly Cys11, abolished H2O2-induced disulfide-linked dimers, and weaken H2O2 damage on So-MntR binding, indicating that H2O2 inactivates So-MntR via disulfide-linked dimerization. So-MntR C123S mutant was extremely sensitive to H2O2 oxidization in dimerization/oligomerization, probably because the mutagenesis caused a conformational change that facilitates Cys11-Cys156 disulfide-linkage. Intermolecular Cys11-Cys11 disulfide was detected in C123S/C156S double mutant. Redox Western blot detected So-MntR oligomers in air-exposed cells, but remarkably decreased upon H2O2 pulsing, suggesting a proteolysis of the disulfide-linked So-MntR oligomers. Remarkably, elevated C11S and C156S but much lower C123S proteins were detected in H2O2-pulsed cells, confirming Cys11 and Cys156 contributed to H2O2-induced oligomerization and degradation. Accordingly, in the C11S and C156S mutants, expression of mntABC and cellular Mn2+ were decreased but H2O2 susceptibility increased. While in the C123S mutant, increased mntABC expression, cellular Mn2+ content and Mn-mediated H2O2 survival were determined. Given the wide distribution of Cys11 in streptococcal DtxR-like metalloregulators, the disclosed redox regulatory function and mechanism of So-MntR can be employed by the DtxR family proteins in bacterial resistance to oxidative stress.

from #AlexandrosSfakianakis via Alexandros G.Sfakianakis on Inoreader http://ift.tt/2l68i8R
via IFTTT