Profiling Protein S‐Sulfination with Maleimide‐Linked Probes

• Published in: Chembiochem : a European journal of chemical biology Vol. 18; no. 20; pp. 2028 - 2032
• Main Authors: Kuo, Yu‐Hsuan, Konopko, Aaron M., Borotto, Nicholas B., Majmudar, Jaimeen D., Haynes, Sarah E., Martin, Brent R.
• Format: Journal Article
• Language: English
• Published: Germany Wiley Subscription Services, Inc 18-10-2017
• Subjects: Alkylation; Cysteine; Electron transport; Electron transport chain; Endoplasmic reticulum; HEK293 Cells; Humans; Lipopolysaccharides; Maleimides - chemistry; Michael addition; Models, Molecular; Molecular Probes - chemistry; Oxidation; Pharmacology; Post-translation; Protein Conformation; protein modifications; Protein S; Proteins; Proteins - chemistry; Proteins - metabolism; redox chemistry; Rodents; Sulfinic Acids - metabolism; sulfur; Sulfur - metabolism; Thiols; Michael addition; sulfur; protein modifications; cysteine; redox chemistry
• ISSN: 1439-4227; 1439-7633
• DOI: 10.1002/cbic.201700137

Cysteine residues are susceptible to oxidation to form S‐sulfinyl (R‐SO2H) and S‐sulfonyl (R‐SO3H) post‐translational modifications. Here we present a simple bioconjugation strategy to label S‐sulfinated proteins by using reporter‐linked maleimides. After alkylation of free thiols with iodoacetamide, S‐sulfinated cysteines react with maleimide to form a sulfone Michael adduct that remains stable under acidic conditions. Using this sequential alkylation strategy, we demonstrate differential S‐sulfination across mouse tissue homogenates, as well as enhanced S‐sulfination following pharmacological induction of endoplasmic reticulum stress, lipopolysaccharide stimulation, and inhibitors of the electron transport chain. Overall, this study reveals a broadened profile of maleimide reactivity across cysteine modifications, and outlines a simple method for profiling the physiological role of cysteine S‐sulfination in disease.
Maleimide, but not iodoacetamide, reacts with aryl and alkyl sulfinic acid standards and S‐sulfinated proteins to give a sulfonyl‐succinimide adduct that is stable under acidic conditions. This sequential alkylation strategy can be used for selective sulfinic acid labeling in biological samples. This study reveals a broadened profile of maleimide reactivity across cysteine modifications in proteins.