Mass spectrometry-based direct detection of multiple types of protein thiol modifications in pancreatic beta cells under endoplasmic reticulum stress.
Redox Biol
; 46: 102111, 2021 10.
Article
em En
| MEDLINE
| ID: mdl-34425387
ABSTRACT
Thiol-based post-translational modifications (PTMs) play a key role in redox-dependent regulation and signaling. Functional cysteine (Cys) sites serve as redox switches, regulated through multiple types of PTMs. Herein, we aim to characterize the complexity of thiol PTMs at the proteome level through the establishment of a direct detection workflow. The LC-MS/MS based workflow allows for simultaneous quantification of protein abundances and multiple types of thiol PTMs. To demonstrate its utility, the workflow was applied to mouse pancreatic ß-cells (ß-TC-6) treated with thapsigargin to induce endoplasmic reticulum (ER) stress. This resulted in the quantification of >9000 proteins and multiple types of thiol PTMs, including intra-peptide disulfide (S-S), S-glutathionylation (SSG), S-sulfinylation (SO2H), S-sulfonylation (SO3H), S-persulfidation (SSH), and S-trisulfidation (SSSH). Proteins with significant changes in abundance were observed to be involved in canonical pathways such as autophagy, unfolded protein response, protein ubiquitination pathway, and EIF2 signaling. Moreover, ~500 Cys sites were observed with one or multiple types of PTMs with SSH and S-S as the predominant types of modifications. In many cases, significant changes in the levels of different PTMs were observed on various enzymes and their active sites, while their protein abundance exhibited little change. These results provide evidence of independent translational and post-translational regulation of enzyme activity. The observed complexity of thiol modifications on the same Cys residues illustrates the challenge in the characterization and interpretation of protein thiol modifications and their functional regulation.
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Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Compostos de Sulfidrila
/
Células Secretoras de Insulina
Tipo de estudo:
Diagnostic_studies
Limite:
Animals
Idioma:
En
Ano de publicação:
2021
Tipo de documento:
Article