Dataset Information

Stochiometric Quantification of the Thiol Redox Proteome Reveals Subcellular Compartmentalization and Susceptibility to Oxidative Perturbations

ABSTRACT: Posttranslational modifications of protein cysteine thiols play a significant role in redox regulation and the pathogenesis of human diseases. However, the cellular redox landscape in terms of quantitative, site-specific occupancies of thiol modifications at the proteome level, especially under physiological conditions, is still largely uncharacterized. Herein, we report the site occupancies of both S-glutathionylation (SSG) and total reversible thiol oxidation (total oxidation) in RAW 264.7 macrophage cells under basal conditions. The occupancies of thiol modifications for ~4,000 cysteine sites were quantified, which revealed a mean site occupancy of 4.0% for SSG and 11.9% for total oxidation, respectively. Correlations between site occupancies and structural features such as pKa, relative residue surface accessibility, and hydrophobicity were observed. Proteome-wide site occupancy analysis also revealed a strong subcellular compartmentalization in thiol redox status, where the average occupancies of SSG and total oxidation in distinct compartments correlate well with the redox potentials of respective organelles.

INSTRUMENT(S): LTQ Orbitrap Velos

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Macrophage Cell Line, Macrophage

SUBMITTER: Matthew Monroe

LAB HEAD: Wei-Jun Qian

PROVIDER: PXD019913 | Pride | 2020-08-13

REPOSITORIES: Pride

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Publications

Stochiometric quantification of the thiol redox proteome of macrophages reveals subcellular compartmentalization and susceptibility to oxidative perturbations.

Duan Jicheng J Zhang Tong T Gaffrey Matthew J MJ Weitz Karl K KK Moore Ronald J RJ Li Xiaolu X Xian Ming M Thrall Brian D BD Qian Wei-Jun WJ

Redox biology 20200721

Posttranslational modifications of protein cysteine thiols play a significant role in redox regulation and the pathogenesis of human diseases. Herein, we report the characterization of the cellular redox landscape in terms of quantitative, site-specific occupancies of both S-glutathionylation (SSG) and total reversible thiol oxidation (total oxidation) in RAW 264.7 macrophage cells under basal conditions. The occupancies of thiol modifications for ~4000 cysteine sites were quantified, revealing ...[more]

PMID: 32750668

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Project description:This proteomic study This proteomic study focuses on the role of reversible thiol oxidation after hexyl aminolevulinate mediated PDT (HAL-PDT). The human epidermoid carcinoma cell line A431 was used as model system and red light as light source, a clinical relevant in vitro model. The light dose dependent cell cytotoxicity was measured by resazurin assay. The most clinical relevant dose, LD100, was used for the proteomic part of the study and cells where harvested 30 min after treatment. The reversibly oxidized thiol proteins were selected by biotinylation and identified by mass spectrometry. This proteomic technique revealed over 1100 thiol proteins which were reversibly oxidized after HAL-PDT. Identified proteins were analysed by bioinformatics (IPA and GO) and a list of reliable identified proteins (282 proteins) where further analysed. Both the cellular location and function was determined for these proteins. In addition, 18 of the proteins where identified as redox regulated, 36 to be a part of the apoptotic pathway and 9 to be both redox regulated and a part of apoptotic pathway. From these results we suggest redox regulated proteins as a trigger mechanism for PDT induced apoptosis.focuses on the role of reversible thiol oxidation after hexyl aminolevulinate mediated PDT (HAL-PDT). The human epidermoid carcinoma cell line A431 was used as model system and red light as light source, a clinical relevant in vitro model. The light dose dependent cell cytotoxicity was measured by resazurin assay. The most clinical relevant dose, LD100, was used for the proteomic part of the study and cells where harvested 30 min after treatment. The reversibly oxidized thiol proteins were selected by biotinylation and identified by mass spectrometry. This proteomic technique revealed over 1100 thiol proteins which were reversibly oxidized after HAL-PDT. Identified proteins were analysed by bioinformatics (IPA and GO) and a list of reliable identified proteins (282 proteins) where further analysed. Both the cellular location and function was determined for these proteins. In addition, 18 of the proteins where identified as redox regulated, 36 to be a part of the apoptotic pathway and 9 to be both redox regulated and a part of apoptotic pathway. From these results we suggest redox regulated proteins as a trigger mechanism for PDT induced apoptosis.

Dataset Information

Stochiometric Quantification of the Thiol Redox Proteome Reveals Subcellular Compartmentalization and Susceptibility to Oxidative Perturbations

Publications

Stochiometric quantification of the thiol redox proteome of macrophages reveals subcellular compartmentalization and susceptibility to oxidative perturbations.

Similar Datasets

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