title: Mitochondrial alpha-ketoglutarate dehydrogenase complex generates reactive oxygen species.
abstract: Mitochondria-produced reactive oxygen species (ROS) are thought to contribute to cell death caused by a multitude of pathological conditions. The molecular sites of mitochondrial ROS production are not well established but are generally thought to be located in complex I and complex III of the electron transport chain. We measured H(2)O(2) production, respiration, and NADPH reduction level in rat brain mitochondria oxidizing a variety of respiratory substrates. Under conditions of maximum respiration induced with either ADP or carbonyl cyanide p-trifluoromethoxyphenylhydrazone,alpha-ketoglutarate supported the highest rate of H(2)O(2) production. In the absence of ADP or in the presence of rotenone, H(2)O(2) production rates correlated with the reduction level of mitochondrial NADPH with various substrates, with the exception of alpha-ketoglutarate. Isolated mitochondrial alpha-ketoglutarate dehydrogenase (KGDHC) and pyruvate dehydrogenase (PDHC) complexes produced superoxide and H(2)O(2). NAD(+) inhibited ROS production by the isolated enzymes and by permeabilized mitochondria. We also measured H(2)O(2) production by brain mitochondria isolated from heterozygous knock-out mice deficient in dihydrolipoyl dehydrogenase (Dld). Although this enzyme is a part of both KGDHC and PDHC, there was greater impairment of KGDHC activity in Dld-deficient mitochondria. These mitochondria also produced significantly less H(2)O(2) than mitochondria isolated from their littermate wild-type mice. The data strongly indicate that KGDHC is a primary site of ROS production in normally functioning mitochondria.
authors.affiliation: Department of Neurology and Neuroscience, Weill Medical College, Cornell University, New York, New York 10021, USA.
authors.forename: Anatoly A
authors.initials: AA
authors.lastname: Starkov
authors.forename: Gary
authors.initials: G
authors.lastname: Fiskum
authors.forename: Christos
authors.initials: C
authors.lastname: Chinopoulos
authors.forename: Beverly J
authors.initials: BJ
authors.lastname: Lorenzo
authors.forename: Susan E
authors.initials: SE
authors.lastname: Browne
authors.forename: Mulchand S
authors.initials: MS
authors.lastname: Patel
authors.forename: M Flint
authors.initials: MF
authors.lastname: Beal
mesh_terms: D000244:Adenosine Diphosphate / Q000494:pharmacology; D000818:Animals; D000968:Antimycin A / Q000031:analogs & derivatives* / Q000494:pharmacology; D003067:Coenzymes; D008058:Dihydrolipoamide Dehydrogenase / Q000172:deficiency / Q000235:genetics / Q000378:metabolism; D004579:Electron Transport / Q000187:drug effects / Q000502:physiology; D042967:Electron Transport Complex I / Q000037:antagonists & inhibitors; D006861:Hydrogen Peroxide / Q000378:metabolism; D007425:Intracellular Membranes / Q000502:physiology; D007655:Ketoglutarate Dehydrogenase Complex / Q000378:metabolism*; D007656:Ketoglutaric Acids / Q000378:metabolism; D008564:Membrane Potentials; D051379:Mice; D018345:Mice, Knockout; D008928:Mitochondria / Q000187:drug effects / Q000201:enzymology*; D009243:NAD / Q000378:metabolism; D009249:NADP / Q000378:metabolism; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D009840:Oligomycins / Q000494:pharmacology; D010084:Oxidation-Reduction; D016548:Prosencephalon / Q000201:enzymology / Q000648:ultrastructure; D011768:Pyruvate Dehydrogenase Complex / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism*; D012402:Rotenone / Q000494:pharmacology; D019802:Succinic Acid / Q000378:metabolism; D013482:Superoxide Dismutase / Q000494:pharmacology; D013481:Superoxides / Q000378:metabolism; D014451:Ubiquinone / Q000031:analogs & derivatives* / Q000032:analysis
chemical_list: D003067:Coenzymes; D007656:Ketoglutaric Acids; D009419:Nerve Tissue Proteins; D009840:Oligomycins; D011768:Pyruvate Dehydrogenase Complex; D017382:Reactive Oxygen Species; D012402:Rotenone; D009243:NAD; D013481:Superoxides; D014451:Ubiquinone; D009249:NADP; D000244:Adenosine Diphosphate; D000968:Antimycin A; C004510:blastmycin; D019802:Succinic Acid; D006861:Hydrogen Peroxide; D013482:Superoxide Dismutase; D007655:Ketoglutarate Dehydrogenase Complex; D008058:Dihydrolipoamide Dehydrogenase; D042967:Electron Transport Complex I; C024989:coenzyme Q10; C030571:ubiquinone 9