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DYn-2 based identification of Arabidopsis sulfenomes

ABSTRACT: Identifying the sulfenylation state (-SOH) of stressed cells. Here, we optimized an in vivo trapping method of sulfenic acids in hydrogen peroxide (H2O2) stressed plant cells. With click chemistry, the dimedone based alkyne functionalized DYn-2 probe was biotinylated for subsequent streptavidin enrichment of sulfenylated proteins. the DYn-2 modification and DYn-2-Biotin-azide modification were unknown modifications, we therefore chose N-D-glucuronylglycine [MOD00067] and N-palmitoyl-S-(sn-1-2,3-dipalmitoyl-glycerol)cysteine [MOD00444]

INSTRUMENT(S): Q Exactive

ORGANISM(S): Arabidopsis Thaliana (mouse-ear Cress)

TISSUE(S): Plant Cell

SUBMITTER: Jarne Pauwels

LAB HEAD: Kris Gevaert

PROVIDER: PXD001562 | Pride | 2016-06-27

REPOSITORIES: Pride

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			Action	DRS
	E03925_2_3704_Salma_notTreated.raw	Raw
	E03927_2_3704_Salma_H2O2Treated.raw	Raw
	E05163_1d_4020_Salma_Akter_with_H2O2.raw	Raw
	E05165_1d_4020_Salma_Akter_without_H2O2.raw	Raw
	E06178_1t_20_4269_SalmaAkter_treated.raw	Raw

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Publications

DYn-2 Based Identification of Arabidopsis Sulfenomes.

Akter Salma S Huang Jingjing J Bodra Nandita N De Smet Barbara B Wahni Khadija K Rombaut Debbie D Pauwels Jarne J Gevaert Kris K Carroll Kate K Van Breusegem Frank F Messens Joris J

Molecular & cellular proteomics : MCP 20150218 5

Identifying the sulfenylation state of stressed cells is emerging as a strategic approach for the detection of key reactive oxygen species signaling proteins. Here, we optimized an in vivo trapping method for cysteine sulfenic acids in hydrogen peroxide (H2O2) stressed plant cells using a dimedone based DYn-2 probe. We demonstrated that DYn-2 specifically detects sulfenylation events in an H2O2 dose- and time-dependent way. With mass spectrometry, we identified 226 sulfenylated proteins after H2 ...[more]

PMID: 25693797

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In situ identification of S-sulfenylation sites in Arabidopsis thaliana using the genetic YAP1 probe

Project description:Hydrogen peroxide (H2O2) reacts, directly or indirectly, with cysteines to form cysteine sulfenic acid, also known as S-sulfenylation. This cysteine oxidation steers diverse cellular processes by altering protein interactions, trafficking, conformation, and function. We present a method to identify S-sulfenylated cysteines sites in proteins using a genetic probe based on the yeast AP-1–like (YAP1) transcription factor that specifically reacts with sulfenic acid sites to form mixed disulfides. After a tryptic digest and with the help of an antibody directed against a 7-amino-acid YAP1C-derived peptide that contains the redox-active cysteine, we enriched for disulfide-linked peptides. The mass spectral characteristics for fragment ions of the mixed disulfides made it possible to identify 1,747 S-sulfenylation protein sites in Arabidopsis under H2O2 stress. Furthermore, cross-study comparison shows that 55% of cross-linked sites match with previously reported S-sulfenylated, S-nitrosylated and reversibly oxidized cysteines in Arabidopsis. These include well-characterized redox-sensitive cysteines, such as Cys20 of DEHYDROASCORBATE REDUCTASE 2 (DHAR2) and Cys181 of MAP KINASE 4 (MAPK4). Altogether, we describe a novel approach to identify S-sulfenylated sites in situ using the YAP1C probe, thereby offering a non-invasive manner to study site-specific cysteine oxidation and which can be applied for identification of S-sulfenylated sites at the organellar level.

2020-12-15 | PXD016723 | Pride

Mining for protein S-sulfenylation in Arabidopsis uncovers redox-sensitive sites

Project description:Hydrogen peroxide (H2O2) is an important messenger molecule for diverse cellular processes. H2O2 oxidizes proteinaceous cysteinyl thiols to sulfenic acid, also known as S-sulfenylation, thereby affecting the protein conformation and functionality. Although many proteins have been identified as S-sulfenylation targets in plants, site-specific mapping and quantification remain largely unexplored. By means of peptide-centric chemoproteomics, 1,537 S-sulfenylated sites were mapped on more than 1,000 proteins in Arabidopsis thaliana cells. The H2O2 sensitivity was quantified of more than 70% of these endogenous oxidation events toward exogenous H2O2 stimulation. Proteins involved in RNA and metabolic processing were identified as hotspots for S-sulfenylation. Moreover, S-sulfenylation frequently occurred on cysteines located in catalytic sites of enzymes or on cysteines involved in metal binding, hinting at direct mode-of-actions for redox regulation. Comparison of human and Arabidopsis S-sulfenylation datasets provided 155 conserved S-sulfenylated cysteines, including Cys181 of the Arabidopsis MITOGEN-ACTIVATED PROTEIN KINASE4 (AtMAPK4) that corresponds to Cys161 in the human MAPK1, which is speculated to be a redox-regulatory site. Replacement of the noncatalytic Cys181 of the recombinant AtMAPK4 by the redox-insensitive serine decreased the protein kinase activity, emphasizing the importance of this noncatalytic cysteine. Altogether, we quantitatively mapped the S-sulfenylated cysteines in Arabidopsis plants under oxidative stress and delivered an unprecedented inventory for unraveling the precise role of these oxidized cysteines in plant redox signaling.

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Sulfenylation of Human Liver and Kidney Microsomal Cytochromes P450 and Other Drug Metaboing Enzymes as a Response to Redox Alteration

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Aerobic and Anaerobic Growth of Streptococcus pneumoniae D39

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2016-12-08 | GSE19791 | GEO

We applied an iodoacetyl tandem mass tags (iodoTMT)-based proteomic approach to analyze the redox proteome of Brassica napus seedlings under control and salt-stressed conditions. We identified 1,821 sulfenylated sites in 912 proteins from all samples.

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