title: Quantitative Persulfide Site Identification (qPerS-SID) Reveals Protein Targets of H2S Releasing Donors in Mammalian Cells.
abstract: H2S is an important signalling molecule involved in diverse biological processes. It mediates the formation of cysteine persulfides (R-S-SH), which affect the activity of target proteins. Like thiols, persulfides show reactivity towards electrophiles and behave similarly to other cysteine modifications in a biotin switch assay. In this manuscript, we report on qPerS-SID a mass spectrometry-based method allowing the isolation of persulfide containing peptides in the mammalian proteome. With this method, we demonstrated that H2S donors differ in their efficacy to induce persulfides in HEK293 cells. Furthermore, data analysis revealed that persulfide formation affects all subcellular compartments and various cellular processes. Negatively charged amino acids appeared more frequently adjacent to cysteines forming persulfides. We confirmed our proteomic data using pyruvate kinase M2 as a model protein and showed that several cysteine residues are prone to persulfide formation finally leading to its inactivation. Taken together, the site-specific identification of persulfides on a proteome scale can help to identify target proteins involved in H2S signalling and enlightens the biology of H2S and its releasing agents.
authors.affiliation: Pharmazentrum Frankfurt/ZAFES, Universitätsklinikum Frankfurt, Frankfurt am Main, Germany.
authors.forename: Sebastian
authors.initials: S
authors.lastname: Longen
authors.affiliation: Functional Proteomics, SFB 815 Core Unit, Goethe University Frankfurt, Frankfurt, Germany.
authors.forename: Florian
authors.initials: F
authors.lastname: Richter
authors.affiliation: Pharmazentrum Frankfurt/ZAFES, Universitätsklinikum Frankfurt, Frankfurt am Main, Germany.
authors.forename: Yvette
authors.initials: Y
authors.lastname: Köhler
authors.affiliation: Functional Proteomics, SFB 815 Core Unit, Goethe University Frankfurt, Frankfurt, Germany.
authors.forename: Ilka
authors.initials: I
authors.lastname: Wittig
authors.affiliation: Pharmazentrum Frankfurt/ZAFES, Universitätsklinikum Frankfurt, Frankfurt am Main, Germany.
authors.forename: Karl-Friedrich
authors.initials: KF
authors.lastname: Beck
authors.affiliation: Pharmazentrum Frankfurt/ZAFES, Universitätsklinikum Frankfurt, Frankfurt am Main, Germany.
authors.forename: Josef
authors.initials: J
authors.lastname: Pfeilschifter
mesh_terms: D000595:Amino Acid Sequence; D002853:Chromatography, Liquid / Q000379:methods; D003545:Cysteine / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism; D004220:Disulfides / Q000737:chemistry / Q000378:metabolism; D057809:HEK293 Cells; D006801:Humans; D006862:Hydrogen Sulfide / Q000737:chemistry / Q000378:metabolism; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism; D020543:Proteome / Q000737:chemistry* / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D013440:Sulfides / Q000737:chemistry* / Q000378:metabolism; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D004220:Disulfides; D010455:Peptides; D020543:Proteome; D013440:Sulfides; C051552:persulfides; C116840:cysteine persulfide; D003545:Cysteine; D006862:Hydrogen Sulfide