Protocol for detecting protein S-glutathionylation in Arabidopsis thaliana under oxidative stress using a non-transgenic chemical toolkit

Protein S-glutathionylation is a redox-sensitive post-translational modification that regulates proteostasis and signaling during oxidative stress. However, its in vivo detection is challenging due to labile disulfide bonds and ex vivo oxidation artifacts. Here, we present a protocol for detecting protein glutathionylation in Arabidopsis thaliana using a non-transgenic toolkit. We describe steps for using propargyl glycine (PG) metabolic labeling and click chemistry. We detail workflows for global quantification, target enrichment for western blotting or liquid chromatography-tandem mass spectrometry (LC-MS/MS), and in situ spatial visualization. For complete details on the development, use and execution of this protocol, please refer to Zhao et al. 1 1. Zhao, S. ∙ Xie, Z. ∙ Chen, X. ... Duet between stress granules and glutathionylation regulates cytosolic redox state to maintain proteostasis in Arabidopsis Mol. Plant. 2026; 19 :606-628 Google Scholar