title: Effects of oxidative stress on protein thiols and disulphides in Mytilus edulis revealed by proteomics: actin and protein disulphide isomerase are redox targets.
abstract: Many proteins contain cysteines which are sensitive to oxidation. This is sometimes reversible through interaction with glutathione, glutaredoxin or thioredoxin systems making these proteins potential sensors of oxidative stress. In this study we analysed whether there was an increase in mixed disulphide bond (-S-S-) formation in the blue mussel Mytilus edulis in response to menadione. This was achieved by initially blocking reduced thiols with N-ethylmaleimide, -S-S- were then reduced with dithiothreitol (DTT) and labelled with 5-iodoacetamidofluorescein (5-IAF). Free -SHs were also labelled directly with 5-IAF. Separations were performed on 1D or 2D SDS PAGE and images analysed. There was an increase in -S-S- in response to menadione and detection of changes in oxidised proteins was easier than that of changes in the amount of reduced proteins. Protein disulphide isomerase (PDI) was labelled both as -SH and -S-S-, underlining its involvement in the redox status of the animal. A glutathione transferase (GST P1-1) forms an inter-chain disulphide bridge in response to menadione.
authors.affiliation: Proteomics Research Group, Department of Biochemistry, Environmental Research Institute, University College Cork, Cork, Ireland.
authors.forename: B
authors.initials: B
authors.lastname: McDonagh
authors.forename: D
authors.initials: D
authors.lastname: Sheehan
mesh_terms: D000199:Actins / Q000378:metabolism*; D000818:Animals; D004220:Disulfides / Q000378:metabolism*; D008958:Models, Molecular; D049880:Mytilus edulis / Q000378:metabolism*; D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000502:physiology*; D019704:Protein Disulfide-Isomerases / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry / Q000378:metabolism*; D040901:Proteomics; D013438:Sulfhydryl Compounds / Q000378:metabolism*
chemical_list: D000199:Actins; D004220:Disulfides; D011506:Proteins; D013438:Sulfhydryl Compounds; D019704:Protein Disulfide-Isomerases