Proteome‐Wide Analysis of Cysteine S‐Sulfenylation Using a Benzothiazine‐Based Probe

Oxidation of a protein cysteinyl thiol (Cys‐SH) to S‐sulfenic acid (Cys‐SOH) by a reactive oxygen species (e.g., hydrogen peroxide), which is termed protein S‐sulfenylation, is a reversible post‐translational modification that plays a crucial role in redox regulation of protein function in various biological processes. Due to its intrinsically labile nature, protein S‐sulfenylation cannot be directly detected or analyzed. Chemoselective probing has been the method of choice for analyzing S‐sulfenylated proteins either in vitro or in situ , as it allows stabilization and direct detection of this transient oxidative intermediate. However, it remains challenging to globally pinpoint the specific S‐sulfenylated cysteine sites on complex proteomes and to quantify their dynamic changes upon oxidative stress. This unit describes how a benzothiazine‐based chemoselective probe called BTD and mass spectrometry based chemoproteomics can be used to globally and site‐specifically identify and quantify protein S‐sulfenylation. © 2018 by John Wiley & Sons, Inc.