Characterization of the enzymes involved in hydrogen sulfide (H2S) detoxification and H2S-reactive nitrogen species (RNS) crosstalk in bacterial pathogens.

摘要

Recent studies suggest that hydrogen sulfide (H2S) functions as a signaling molecule in mammals, but can also protect bacteria from the effects of antibiotic stress. The role of cellular H2S homeostasis and reactive sulfur species (RSS) in bacterial physiology is poorly understood. We discovered and characterized the cst operon from the major human pathogen Staphylococcus aureus ( S. aureus; Sau), which is induced by cellular H2S stress via direct reaction of the transcriptional repressor CstR with low molecular weight (LMW) persulfides and inorganic polysulfides. In this work, the enzymes encoded by two genes of the cst operon, SQR and CstB, have been biochemically characterized and shown to comprise the core of a sulfide oxidation system analogous to the human mitochondrial H2S detoxification system. SQR is a flavin adenine dinucleotide (FAD)-dependent enzyme that catalyzes the initial H2S oxidation to sulfane sulfur (S0) which reacts with cellular LMW thiols to form LMW thiol persulfides. These LMW persulfides are further oxidized by CstB, a non-heme Fe(II)-containing multidomain persulfide dioxygenase-sulfurtransferase, to thiosulfate and reduced LMW thiols. We propose that S. aureus utilizes this H 2S oxidation system to avoid the highly cytotoxic metabolite sulfite through an intramolecular domain-domain interaction in CstB, as well as the accumulation of LMW persulfides that form in the presence of H2S. We also discovered and preliminarily characterized a cst-like operon from another major human pathogen Enterococcus faecalis (E. faecalis; Ef) that also responds to cellular H2 S stress. This operon encodes a CoADR-RHD fusion protein that we show reduces CoA persulfide to CoA and H2S, whose fate is not yet known. Complementation of a S. aureus CstB deletion strain with the E. faecalis CoADR-RHD reverses the sulfide-induced growth phenotype of the DeltacstB strain. This suggests that these systems have evolved to protect the coenzyme A pool against persulfidation under conditions of high intracellular RSS.;Previous work has shown that upregulation of the endogenous H2S level and bacterially synthesized nitric oxide (NO) protects S. aureus from the effects of antibiotic stress via a mechanism we propose involves the intermediacy of nitroxyl (HNO). We showed that exogenous HNO stress directly induces both cst operon from S. aureus and cst-like operon from E. faecalis, probably via the upregulation of cellular H2S levels, which has been shown to increase the cellular concentrations of LMW persulfides. The implications of these findings are discussed.