Abstract
S-Nitrosothiols (RSNOs) represent an important class of post-translational modifications that preserve and amplify the actions of nitric oxide and regulate enzyme activity. Several regulatory proteins are now verified targets of cellular S-nitrosation, and the direct detection of S-nitrosated residues in proteins has become essential to better understand RSNO-mediated signaling. Current RSNO detection depends on indirect assays that limit their overall specificity and reliability. Herein, we report the reaction of S-nitrosated cysteine, glutathione, and a mutated C165S alkyl hydroperoxide reductase with the water-soluble phosphine tris(4,6-dimethyl-3-sulfonatophenyl)phosphine trisodium salt hydrate (TXPTS). A combination of NMR and MS techniques reveals that these reactions produce covalent S-alkylphosphonium ion adducts (with S-P(+) connectivity), TXPTS oxide, and a TXPTS-derived aza-ylide. Mechanistically, this reaction may proceed through an S-substituted aza-ylide or the direct displacement of nitroxyl from the RSNO group. This work provides a new means for detecting and quantifying S-nitrosated species in solution and suggests that phosphines may be useful tools for understanding the complex physiological roles of S-nitrosation and its implications in cell signaling and homeostasis.