title: A chemical approach for detecting sulfenic acid-modified proteins in living cells.
abstract: Oxidation of the thiol functional group in cysteine (Cys-SH) to sulfenic (Cys-SOH), sulfinic (Cys-SO2H) and sulfonic acids (Cys-SO3H) is emerging as an important post-translational modification that can activate or deactivate the function of many proteins. Changes in thiol oxidation state have been implicated in a wide variety of cellular processes and correlate with disease states but are difficult to monitor in a physiological setting because of a lack of experimental tools. Here, we describe a method that enables live cell labeling of sulfenic acid-modified proteins. For this approach, we have synthesized the probe DAz-1, which is chemically selective for sulfenic acids and cell permeable. In addition, DAz-1 contains an azide chemical handle that can be selectively detected with phosphine reagents via the Staudinger ligation for identification, enrichment and visualization of modified proteins. Through a combination of biochemical, mass spectrometry and immunoblot approaches we characterize the reactivity of DAz-1 and highlight its utility for detecting protein sulfenic acids directly in mammalian cells. This novel method to isolate and identify sulfenic acid-modified proteins should be of widespread utility for elucidating signaling pathways and regulatory mechanisms that involve oxidation of cysteine residues.
authors.affiliation: Life Sciences Institute, University of Michigan, 210 Washtenaw Avenue, Ann Arbor, Michigan 48109-2216, USA.
authors.forename: Khalilah G
authors.initials: KG
authors.lastname: Reddie
authors.forename: Young Ho
authors.initials: YH
authors.lastname: Seo
authors.forename: Wilson B
authors.initials: WB
authors.lastname: Muse Iii
authors.forename: Stephen E
authors.initials: SE
authors.lastname: Leonard
authors.forename: Kate S
authors.initials: KS
authors.lastname: Carroll
mesh_terms: D000426:Alcohol Dehydrogenase / Q000737:chemistry; D007611:Aprotinin / Q000737:chemistry; D001386:Azides / Q000138:chemical synthesis* / Q000737:chemistry*; D015153:Blotting, Western; D002470:Cell Survival; D002478:Cells, Cultured; D006801:Humans; D019169:Jurkat Cells; D008024:Ligands; D015394:Molecular Structure; D010088:Oxidoreductases / Q000737:chemistry; D012709:Serum Albumin / Q000032:analysis* / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization; D013194:Staining and Labeling; D013434:Sulfenic Acids / Q000737:chemistry*; D013879:Thioredoxins / Q000737:chemistry; D001614:beta-Amylase / Q000737:chemistry
chemical_list: D001386:Azides; C533869:DAz-1 compound; D008024:Ligands; D012709:Serum Albumin; D013434:Sulfenic Acids; D013879:Thioredoxins; D007611:Aprotinin; D010088:Oxidoreductases; D000426:Alcohol Dehydrogenase; C052449:3'-phosphoadenylyl-5'-phosphosulfate reductase; D001614:beta-Amylase