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S-sulfenylation site detection using YAP1C

S-sulfenylation in Arabidopsis thaliana

1747 modifications in 1735 peptides, found in 2560 proteins

Experiment Details

Exp 110


Experimental Setup
TissueCell cultures
Condition30 min 20 mM H2O2
PTM EnrichmentAntibody enriching YAP1C disulfide cross-linked peptides
MS InstrumentQ Exactive
MS/MS Search Parameters
Protein DatabaseAraport11
Decoy StrategyTarget-decoy
FDR Threshold0.01
Search Algorithm(s)pLink 2.3.8
Precursor Mass Tolerance20 ppm
ProteaseTrypsin
Variable ModificationsCarbamidomethyl (C)
Protein N-terminal Acetylation
Oxidation (M)
Other Information
CommentsYAP1C Cys598 disulfide cross-linked peptides were identified by pLink2. Only the Arabidopsis protein linked peptide to YAPC1 CSEIWDR was displayed for correct display in Plant PTM Viewer.


Publication Information

Wei et al., 2020

PubMed ID: 32714340

ProteomeXchange: PXD016723

Abstract

Front Plant Sci. 2020 Jul 2;11:777. doi: 10.3389/fpls.2020.00777. eCollection 
2020.

Identification of Sulfenylated Cysteines in Arabidopsis thaliana Proteins Using 
a Disulfide-Linked Peptide Reporter.

Wei B(1)(2)(3)(4)(5), Willems P(1)(2), Huang J(1)(2), Tian C(6), Yang J(6), 
Messens J(3)(4)(5), Van Breusegem F(1)(2).

Author information:
(1)Department of Plant Biotechnology and Bioinformatics, Ghent University, 
Ghent, Belgium.
(2)VIB Center for Plant Systems Biology, Ghent, Belgium.
(3)VIB-VUB Center for Structural Biology, Brussels, Belgium.
(4)Brussels Center for Redox Biology, Brussels, Belgium.
(5)Structural Biology Brussels, Vrije Universiteit Brussel, Brussels, Belgium.
(6)State Key Laboratory of Proteomics, Beijing Proteome Research Center, 
National Center for Protein Sciences, Beijing Institute of Lifeomics, Beijing, 
China.

In proteins, hydrogen peroxide (H2O2) reacts with redox-sensitive cysteines to 
form cysteine sulfenic acid, also known as S-sulfenylation. These cysteine 
oxidation events can steer diverse cellular processes by altering protein 
interactions, trafficking, conformation, and function. Previously, we had 
identified S-sulfenylated proteins by using a tagged proteinaceous probe based 
on the yeast AP-1-like (Yap1) transcription factor that specifically reacts with 
sulfenic acids and traps them through a mixed disulfide bond. However, the 
identity of the S-sulfenylated amino acid residues within a protein remained 
enigmatic. By using the same transgenic YAP1C probe, we present here a 
technological advancement to identify in situ sulfenylated cysteine sites in 
Arabidopsis thaliana cells under control condition and oxidative stress. 
Briefly, the total extract of transgenic YAP1C A. thaliana cells was initially 
purified on IgG-Sepharose beads, followed by a tryptic digest. Then, the mixed 
disulfide-linked peptides were further enriched at the peptide level on an 
anti-YAP1C-derived peptide (C598SEIWDR) antibody. Subsequent mass spectrometry 
analysis with pLink 2 identified 1,745 YAP1C cross-linked peptides, indicating 
sulfenylated cysteines in over 1,000 proteins. Approximately 55% of these 
YAP1C-linked cysteines had previously been reported as redox-sensitive cysteines 
(S-sulfenylation, S-nitrosylation, and reversibly oxidized cysteines). The 
presented methodology provides a noninvasive approach to identify sulfenylated 
cysteines in any species that can be genetically modified.

Copyright © 2020 Wei, Willems, Huang, Tian, Yang, Messens and Van Breusegem.

DOI: 10.3389/fpls.2020.00777
PMCID: PMC7343964
PMID: 32714340