Proteome-wide Quantification and Characterization of Oxidation-Sensitive Cysteines in Pathogenic Bacteria

Thiol-group oxidation of active and allosteric cysteines is a widespread regulatory posttranslational protein modification. Pathogenic bacteria, including Pseudomonas aeruginosa and Staphylococcus aureus, use regulatory cysteine oxidation to respond to and overcome reactive oxygen species (ROS) encountered in the host environment. To obtain a proteome-wide view of oxidation-sensitive cysteines in these two pathogens, we employed a competitive activity-based protein profiling approach to globally quantify hydrogen peroxide (H2O2) reactivity with cysteines across bacterial proteomes. We identified ∼200 proteins containing H2O2-sensitive cysteines, including metabolic enzymes, transcription factors, and uncharacterized proteins. Additional biochemical and genetic studies identified an oxidation-responsive cysteine in the master quorum-sensing regulator LasR and redox-regulated activities for acetaldehyde dehydrogenase ExaC, arginine deiminase ArcA, and glyceraldehyde 3-phosphate dehydrogenase. Taken together, our data indicate that pathogenic bacteria exhibit a complex, multilayered response to ROS that includes the rapid adaption of metabolic pathways to oxidative-stress challenge. ► Oxidation-sensitive cysteines across P. aeruginosa and S. aureus proteomes profiled ► P. aeruginosa quorum-sensing regulator LasR senses oxidative stress via Cys79 ► Oxidative stress inhibits bacterial metabolic enzymes via catalytic cysteine oxidation ► Arginine deiminase activation by Cys oxidation confers resistance to phagocyte killing