Dataset Information

Quantification of thioether-linked glutathione modifications in human lens proteins.

ABSTRACT: Dehydroalanine (DHA) and dehydrobutyrine (DHB) intermediates, formed through ?-elimination, induce protein irreversible glutathionylation and protein-protein crosslinking in human lens fiber cells. In total, irreversible glutathionylation was detected on 52 sites including cysteine, serine and threonine residues in 18 proteins in human lenses. In this study, the levels of GSH modification on three serine residues and four cysteine residues located in seven different lens proteins isolated from different regions and different aged lenses were quantified. The relative levels of modification (modified/nonmodified) were site-specific and age-related, ranging from less than 0.05% to about 500%. The levels of modification on all of the sites quantified in the lens cortex increased with age and GSH modification also increased from cortex to outer nucleus region suggesting an age-related increase of modification. The levels of modification on sites located in stable regions of the proteins such as Cys117 of ?A3, Cys80 of ?B1 and Cys27 of ?S, continued increasing in inner nucleus, but modification on sites located in regions undergoing degradation with age decreased in the inner nucleus suggesting GSH modified proteins were more susceptible to further modification. Irreversible GSH modification in cataract lenses was typically higher than in age-matched normal lenses, but the difference did not reach statistical significance for a majority of sites, with the exception Cys117 of ?A3 crystallin in WSF. Except for S59 of ?A and ?B crystallins, GSH modification did not induce protein insolubility suggesting a possible role for this modification in protection from protein-protein crosslinking.

SUBMITTER: Wang Z

PROVIDER: S-EPMC6167147 | biostudies-literature | 2018 Oct

REPOSITORIES: biostudies-literature

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Quantification of thioether-linked glutathione modifications in human lens proteins.

Wang Zhen Z Schey Kevin L KL

Experimental eye research 20180604

Dehydroalanine (DHA) and dehydrobutyrine (DHB) intermediates, formed through β-elimination, induce protein irreversible glutathionylation and protein-protein crosslinking in human lens fiber cells. In total, irreversible glutathionylation was detected on 52 sites including cysteine, serine and threonine residues in 18 proteins in human lenses. In this study, the levels of GSH modification on three serine residues and four cysteine residues located in seven different lens proteins isolated from d ...[more]

PMID: 29879394

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Project description:PURPOSE:To determine global protein expression changes in the lens of the GSH-deficient LEGSKO mouse model of age-related cataract for comparison with recently published gene expression data obtained by RNA-Seq transcriptome analysis. METHODS:Lenses were separated into epithelial and cortical fiber sections, digested with trypsin, and labeled with isobaric tags (10-plex TMTTM). Peptides were analyzed by LC-MS/MS (Orbitrap Fusion) and mapped to the mouse proteome for relative protein quantification. RESULTS:1871 proteins in lens epithelia and 870 proteins in lens fiber cells were quantified. 40 proteins in LEGSKO epithelia, 14 proteins in LEGSKO fiber cells, 22 proteins in buthionine sulfoximine (BSO)-treated LEGSKO epithelia, and 55 proteins in BSO-treated LEGSKO fiber cells had significantly (p<0.05, FDR<0.1) altered protein expression compared to WT controls. HSF4 and MAF transcription factors were the most common upstream regulators of the response to GSH-deficiency. Many detoxification proteins, including aldehyde dehydrogenases, peroxiredoxins, and quinone oxidoreductase, were upregulated but several glutathione S-transferases were downregulated. Several cellular stress response proteins showed regulation changes, including an upregulation of HERPUD1, downregulation of heme oxygenase, and mixed changes in heat shock proteins. NRF2-regulated proteins showed broad upregulation in BSO-treated LEGSKO fiber cells, but not in other groups. Strong trends were seen in downregulation of lens specific proteins, including ?- and ?-crystallins, lengsin, and phakinin, and in epithelial-mesenchymal transition (EMT)-related changes. Western blot analysis of LEGSKO lens epithelia confirmed expression changes in several proteins. CONCLUSIONS:This dataset confirms at the proteomic level many findings from the recently determined GSH-deficient lens transcriptome and provides new insight into the roles of GSH in the lens, how the lens adapts to oxidative stress, and how GSH affects EMT in the lens.

Dataset Information

Quantification of thioether-linked glutathione modifications in human lens proteins.

Publications

Quantification of thioether-linked glutathione modifications in human lens proteins.

Similar Datasets

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