title: Harnessing Redox Cross-Reactivity To Profile Distinct Cysteine Modifications.
abstract: Cysteine S-nitrosation and S-sulfination are naturally occurring post-translational modifications (PTMs) on proteins induced by physiological signals and redox stress. Here we demonstrate that sulfinic acids and nitrosothiols react to form a stable thiosulfonate bond, and leverage this reactivity using sulfinate-linked probes to enrich and annotate hundreds of endogenous S-nitrosated proteins. In physiological buffers, sulfinic acids do not react with iodoacetamide or disulfides, enabling selective alkylation of free thiols and site-specific analysis of S-nitrosation. In parallel, S-nitrosothiol-linked probes enable enrichment and detection of endogenous S-sulfinated proteins, confirming that a single sulfinic acid can react with a nitrosothiol to form a thiosulfonate linkage. Using this approach, we find that hydrogen peroxide addition increases S-sulfination of human DJ-1 (PARK7) at Cys106, whereas Cys46 and Cys53 are fully oxidized to sulfonic acids. Comparative gel-based analysis of different mouse tissues reveals distinct profiles for both S-nitrosation and S-sulfination. Quantitative proteomic analysis demonstrates that both S-nitrosation and S-sulfination are widespread, yet exhibit enhanced occupancy on select proteins, including thioredoxin, peroxiredoxins, and other validated redox active proteins. Overall, we present a direct, bidirectional method to profile select redox cysteine modifications based on the unique nucleophilicity of sulfinic acids.
authors.affiliation: Department of Chemistry, ‡Program in Chemical Biology, University of Michigan , 930 North University Avenue, Ann Arbor, Michigan 48109, United States.
authors.forename: Jaimeen D
authors.initials: JD
authors.lastname: Majmudar
authors.affiliation: Department of Chemistry, ‡Program in Chemical Biology, University of Michigan , 930 North University Avenue, Ann Arbor, Michigan 48109, United States.
authors.forename: Aaron M
authors.initials: AM
authors.lastname: Konopko
authors.affiliation: Department of Chemistry, ‡Program in Chemical Biology, University of Michigan , 930 North University Avenue, Ann Arbor, Michigan 48109, United States.
authors.forename: Kristin J
authors.initials: KJ
authors.lastname: Labby
authors.affiliation: Department of Chemistry, ‡Program in Chemical Biology, University of Michigan , 930 North University Avenue, Ann Arbor, Michigan 48109, United States.
authors.forename: Christopher T M B
authors.initials: CT
authors.lastname: Tom
authors.affiliation: Department of Chemistry, ‡Program in Chemical Biology, University of Michigan , 930 North University Avenue, Ann Arbor, Michigan 48109, United States.
authors.forename: John E
authors.initials: JE
authors.lastname: Crellin
authors.affiliation: Department of Chemistry, ‡Program in Chemical Biology, University of Michigan , 930 North University Avenue, Ann Arbor, Michigan 48109, United States.
authors.forename: Ashesh
authors.initials: A
authors.lastname: Prakash
authors.affiliation: Department of Chemistry, ‡Program in Chemical Biology, University of Michigan , 930 North University Avenue, Ann Arbor, Michigan 48109, United States.
authors.forename: Brent R
authors.initials: BR
authors.lastname: Martin
mesh_terms: D003429:Cross Reactions; D003545:Cysteine / Q000737:chemistry*; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000737:chemistry; D009603:Nitroso Compounds / Q000737:chemistry; D015513:Oncogene Proteins / Q000737:chemistry; D010084:Oxidation-Reduction; D000071617:Protein Deglycase DJ-1; D013438:Sulfhydryl Compounds / Q000737:chemistry; D013441:Sulfinic Acids / Q000737:chemistry
chemical_list: D047908:Intracellular Signaling Peptides and Proteins; D009603:Nitroso Compounds; D015513:Oncogene Proteins; D013438:Sulfhydryl Compounds; D013441:Sulfinic Acids; C105131:PARK7 protein, human; D000071617:Protein Deglycase DJ-1; D003545:Cysteine