Modulation of mitochondrial complex I activity by reversible Ca2+ and NADH mediated superoxide anion dependent inhibition.
Complex I, a key component of the mitochondrial respiratory chain, exhibits diminished activity as a result of cardiac ischemia/reperfusion. Cardiac ischemia/reperfusion is associated with increases in the levels of mitochondrial Ca(2+) and pro-oxidants. In the current in vitro study, we sought evidence for a mechanistic link between Ca(2+), pro-oxidants, and inhibition of complex I utilizing mitochondria isolated from rat heart. Our results indicate that addition of Ca(2+) to solubilized mitochondria results in loss in complex I activity. Ca(2+) induced a maximum decrease in complex I activity of approximately 35% at low micromolar concentrations over a narrow physiologically relevant pH range. Loss in activity required reducing equivalents in the form of NADH and was not reversed upon addition of EGTA. The antioxidants N-acetylcysteine and superoxide dismutase, but not catalase, prevented inhibition, indicating the involvement of superoxide anion (O2(*-)) in the inactivation process. Importantly, the sulfhydryl reducing agent DTT was capable of fully restoring complex I activity implicating the formation of sulfenic acid and/or disulfide derivatives of cysteine in the inactivation process. Finally, complex I can reactivate endogenously upon Ca(2+) removal if NADH is present and the enzyme is allowed to turnover catalytically. Thus, the present study provides a mechanistic link between three alterations known to occur during cardiac ischemia/reperfusion, mitochondrial Ca(2+) accumulation, free radical production, and complex I inhibition. The reversibility of these processes suggests redox regulation of Ca(2+) handling.
Sadek
Hesham A
HA
Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio 44106-4970, USA.
Szweda
Pamela A
PA
Szweda
Luke I
LI
D000111:Acetylcysteine / Q000494:pharmacology; D000818:Animals; D000838:Anions; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D002374:Catalase / Q000378:metabolism; D003902:Detergents / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000494:pharmacology; D042967:Electron Transport Complex I / Q000378:metabolism*; D005609:Free Radicals; D006863:Hydrogen-Ion Concentration; D008297:Male; D008928:Mitochondria / Q000378:metabolism; D009243:NAD / Q000737:chemistry / Q000378:metabolism*; D016877:Oxidants / Q000378:metabolism; D010084:Oxidation-Reduction; D051381:Rats; D017207:Rats, Sprague-Dawley; D015427:Reperfusion Injury; D013482:Superoxide Dismutase / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism*; D013997:Time Factors
D000838:Anions; D003902:Detergents; D005609:Free Radicals; D016877:Oxidants; D009243:NAD; D013481:Superoxides; D004533:Egtazic Acid; D002374:Catalase; D013482:Superoxide Dismutase; D042967:Electron Transport Complex I; D002118:Calcium; D004229:Dithiothreitol; D000111:Acetylcysteine