Chemical Proteomics Reveals New Targets of Cysteine Sulfinic Acid Reductase

Cysteine sulfinic acid or S-sulfinylation is an oxidative post-translational modification (OxiPTM) that is known to be involved in redox-dependent regulation of protein function, but has been historically difficult to analyze biochemically. To facilitate the detection of S-sulfinylated proteins, we demonstrate that a clickable, electrophilic diazene probe (DiaAlk) enables capture and site-centric proteomic analysis of this OxiPTM. Using this workflow, we revealed a striking difference between sulfenic acid (S-sulfenylation) and S-sulfinylation dynamic response to oxidative stress, indicative of different roles for these OxiPTMs in redox regulation. We also identified >55 heretofore unknown protein substrates of the cysteine sulfinic acid reductase, sulfiredoxin (SRX), extending its function well beyond 2-Cys peroxiredoxins (2-Cys PRDX1–4), and offering new insight into the role of this unique oxidoreductase as a central mediator of ROS-associated diseases, particularly cancer. DiaAlk therefore provides a novel tool to profile S-sulfinylated proteins and study their regulatory mechanisms in cells.