Endogenous Sulfane Sulfur Mediates the Oxidative Stress Response Process in Pseudomonas aeruginosa

Sulfane sulfur species are increasingly recognized as integral cellular components involved in signaling pathways and cytoprotection against oxidative stress in mammals. While their production in bacteria has been extensively studied, their functional role in bacterial oxidative stress defense remains poorly understood. Here, we demonstrate that sulfane sulfur generated by sulfide: quinone oxidoreductase decreases H₂O₂ sensitivity in Pseudomonas aeruginosa PAO1. Notably, this protective mechanism does not depend on sulfane sulfur acting as a direct H₂O₂ scavenger via nucleophilic reactions. Through persulfidation proteomic profiling, we reveal that persulfidation is a prominent post-translational modification in P. aeruginosa, reflecting the prevalence of deprotonated sulfane sulfur species. These species modify cysteine residues in proteins, including the well-known oxidative stress regulator OxyR. Specifically, sulfane sulfur modifies OxyR at Cys199 to form persulfidated OxyR C199-SSH, contributing to a single-Cys activated state that modulates promoter activity and DNA-binding affinity. Furthermore, sulfane sulfur-mediated persulfidation protects the critical cysteine residue of LpdG, a ROS-vulnerable dihydrolipoamide dehydrogenase, from irreversible overoxidation. Although LpdG is not part of the canonical H₂O₂-scavenging system, its preservation is essential for cell viability under oxidative stress. These findings establish endogenous sulfane sulfur species as key mediators of antioxidant defense in P. aeruginosa.