First proteomic study of S-glutathionylation in cyanobacteria

J Proteome Res. 2015 Jan 2;14(1):59-71. doi: 10.1021/pr500625a. Epub 2014 Sep 19.

Abstract

Glutathionylation, the reversible post-translational formation of a mixed disulfide between a cysteine residue and glutathione (GSH), is a crucial mechanism for signal transduction and regulation of protein function. Until now this reversible redox modification was studied mainly in eukaryotic cells. Here we report a large-scale proteomic analysis of glutathionylation in a photosynthetic prokaryote, the model cyanobacterium Synechocystis sp. PCC6803. Treatment of acellular extracts with N,N-biotinyl glutathione disulfide (BioGSSG) induced glutathionylation of numerous proteins, which were subsequently isolated by affinity chromatography on streptavidin columns and identified by nano LC-MS/MS analysis. Potential sites of glutathionylation were also determined for 125 proteins following tryptic cleavage, streptavidin-affinity purification, and mass spectrometry analysis. Taken together the two approaches allowed the identification of 383 glutathionylatable proteins that participate in a wide range of cellular processes and metabolic pathways such as carbon and nitrogen metabolisms, cell division, stress responses, and H2 production. In addition, the glutathionylation of two putative targets, namely, peroxiredoxin (Sll1621) involved in oxidative stress tolerance and 3-phosphoglycerate dehydrogenase (Sll1908) acting on amino acids metabolism, was confirmed by biochemical studies on the purified recombinant proteins. These results suggest that glutathionylation constitutes a major mechanism of global regulation of the cyanobacterial metabolism under oxidative stress conditions.

Keywords: Synechocystis; carbon metabolism; cyanobacteria; cysteine modification; glutathionylation; mixed-disulfide bridge; nitrogen metabolism; proteomics; redox regulation.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Bacterial Proteins / metabolism*
  • Glutathione Disulfide / metabolism*
  • Molecular Sequence Data
  • Peptide Fragments / chemistry
  • Protein Processing, Post-Translational
  • Proteome / metabolism*
  • Proteomics
  • Synechocystis / metabolism*

Substances

  • Bacterial Proteins
  • Peptide Fragments
  • Proteome
  • Glutathione Disulfide