Technical documentation
Clickable Glutathione-Based Identification of Cysteine Glutathionylation
Current protocols, v 3(10), e907
Oct 2023
PMID: 37818879
Featured in Collection : UN Sustainable Development Goals @ Drexel
Abstract
Clickable glutathione is a glutathione-derived chemical probe designed to identify and analyze protein S-glutathionylation, a major cysteine oxidation in redox signaling. An engineered glutathione synthetase mutant (GS M4) is used to synthesize clickable glutathione in cells or in vitro, which affords utility via click chemistry to detect, identify, and quantify glutathionylation on individual or global proteins in biochemical and mass spectrometric analyses. The clickable glutathione approach is valuable for the unequivocal identification of glutathionylated cysteines, among many reversible cysteine oxoforms, via the direct enrichment and detection of glutathionylated proteins or peptides. Clickable glutathione, in combination with GS M4, has demonstrated utility in the mass-spectrometry-based discovery and profiling of new proteins and cysteines for glutathionylation in cell lines in response to physiologic and oxidative stress. The approach is versatile and applicable to validating the glutathionylation of proteins and cysteines in other biochemical analysis beside mass spectrometry. Here, we describe the applications of clickable glutathione and provide detailed protocols for the identification, profiling, and detection of glutathionylated proteins and cysteines. © 2023 Wiley Periodicals LLC. Basic Protocol 1: Identification of glutathionylated cysteine in individual proteins in vitro Basic Protocol 2: Proteomic identification and quantification of glutathionylation Basic Protocol 3: Biochemical validation of glutathionylation in cells.
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Details
- Title
- Clickable Glutathione-Based Identification of Cysteine Glutathionylation
- Creators
- Nadee N J Matarage Don - Drexel UniversityDhanushika S K Kukulage - Drexel UniversityYoung-Hoon Ahn - Drexel University, Chemistry
- Publication Details
- Current protocols, v 3(10), e907
- Publisher
- Wiley
- Grant note
- R01 GM143214 / NIGMS NIH HHS R01 HL131740 / NHLBI NIH HHS
- Resource Type
- Technical documentation
- Language
- English
- Academic Unit
- Chemistry
- Web of Science ID
- WOS:001282193900009
- Scopus ID
- 2-s2.0-85173632104
- Other Identifier
- 991022024011204721
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- Biochemical Research Methods