title: Mitochondrial complex II in the post-ischemic heart: oxidative injury and the role of protein S-glutathionylation.
abstract: Mitochondrial superoxide (O2.) is an important mediator of ischemia/reperfusion (I/R) injury. The O2. generated in mitochondria also acts as a redox signal triggering cellular apoptosis. The enzyme succinate ubiquinone reductase (SQR or complex II) is one of the major mitochondrial components hosting regulatory thiols. Here the intrinsic protein S-glutathionylation (PrSSG) at the 70-kDa FAD-binding subunit of SQR was detected in rat heart and in isolated SQR using an anti-GSH monoclonal antibody. When rats were subjected to 30 min of coronary ligation followed by 24 h of reperfusion, the electron transfer activity (ETA) of SQR in post-ischemic myocardium was significantly decreased by 41.5 +/- 2.9%. The PrSSGs of SQR-70 kDa were partially or completely eliminated in post-ischemic myocardium obtained from in vivo regional I/R hearts or isolated global I/R hearts, respectively. These results were further confirmed by using isolated succinate cytochrome c reductase (complex II + complex III). In the presence of succinate, O2. was generated and oxidized the SQR portion of SCR, leading to a 60-70% decrease in its ETA. The gel band of the S-glutathionylated SQR 70-kDa polypeptide was cut out and digested with trypsin, and the digests were subjected to liquid chromatography/tandem mass spectrometry analysis. One cysteine residue, Cys(90), was involved in S-glutathionylation. These results indicate that the glutathione-binding domain, (77)AAFGLSEAGFNTACVTK(93) (where underline indicates Cys(90)), is susceptible to redox change induced by oxidative stress. Furthermore, in vitro S-glutathionylation of purified SQR resulted in enhanced SQR-derived electron transfer efficiency and decreased formation of the 70-kDa-derived protein thiyl radical induced by O2. . Thus, the decreasing S-glutathionylation and ETA in mitochondrial complex II are marked during myocardial ischemia/reperfusion. This redox-triggered impairment of complex II occurs in the post-ischemic heart and should be useful to identify disease pathogenesis related to reactive oxygen species-induced mitochondrial dysfunction.
authors.affiliation: Division of Cardiovascular Medicine, Department of Internal Medicine, Davis Heart and Lung Research Institute, College of Medicine, The Ohio State University, 473 W. 12th Avenue, Columbus, OH 43210, USA. yeong-renn.chen@osumc.edu
authors.forename: Yeong-Renn
authors.initials: YR
authors.lastname: Chen
authors.forename: Chwen-Lih
authors.initials: CL
authors.lastname: Chen
authors.forename: Douglas R
authors.initials: DR
authors.lastname: Pfeiffer
authors.forename: Jay L
authors.initials: JL
authors.lastname: Zweier
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D004578:Electron Spin Resonance Spectroscopy; D042963:Electron Transport Complex II / Q000378:metabolism*; D005415:Flavins / Q000378:metabolism; D005609:Free Radicals / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism*; D008929:Mitochondria, Heart / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017202:Myocardial Ischemia / Q000378:metabolism*; D018384:Oxidative Stress; D011485:Protein Binding; D021122:Protein Subunits / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D013481:Superoxides / Q000378:metabolism; D053719:Tandem Mass Spectrometry
chemical_list: D005415:Flavins; D005609:Free Radicals; D021122:Protein Subunits; D013481:Superoxides; D042963:Electron Transport Complex II; D005978:Glutathione