title: Redox-dependent modulation of aconitase activity in intact mitochondria.
abstract: It has previously been reported that exposure of purified mitochondrial or cytoplasmic aconitase to superoxide (O(2)(-)(*) or hydrogen peroxide (H(2)O(2)) leads to release of the Fe-alpha from the enzyme's [4Fe-4S](2+) cluster and to inactivation. Nevertheless, little is known regarding the response of aconitase to pro-oxidants within intact mitochondria. In the present study, we provide evidence that aconitase is rapidly inactivated and subsequently reactivated when isolated cardiac mitochondria are treated with H(2)O(2). Reactivation of the enzyme is dependent on the presence of the enzyme's substrate, citrate. EPR spectroscopic analysis indicates that enzyme inactivation precedes release of the labile Fe-alpha from the enzyme's [4Fe-4S](2+) cluster. In addition, as judged by isoelectric focusing gel electrophoresis, the relative level of Fe-alpha release and cluster disassembly does not reflect the magnitude of enzyme inactivation. These observations suggest that some form of posttranslational modification of aconitase other than release of iron is responsible for enzyme inactivation. In support of this conclusion, H(2)O(2) does not exert its inhibitory effects by acting directly on the enzyme, rather inactivation appears to result from interaction(s) between aconitase and a mitochondrial membrane component responsive to H(2)O(2). Nevertheless, prolonged exposure of mitochondria to steady-state levels of H(2)O(2) or O(2)(-)(*) results in disassembly of the [4Fe-4S](2+) cluster, carbonylation, and protein degradation. Thus, depending on the pro-oxidant species, the level and duration of the oxidative stress, and the metabolic state of the mitochondria, aconitase may undergo reversible modulation in activity or progress to [4Fe-4S](2+) cluster disassembly and proteolytic degradation.
authors.affiliation: Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio 44106-4970, USA.
authors.forename: Anne-Laure
authors.initials: AL
authors.lastname: Bulteau
authors.forename: Masao
authors.initials: M
authors.lastname: Ikeda-Saito
authors.forename: Luke I
authors.initials: LI
authors.lastname: Szweda
mesh_terms: D000154:Aconitate Hydratase / Q000037:antagonists & inhibitors / Q000737:chemistry* / Q000378:metabolism*; D000818:Animals; D010450:Endopeptidases / Q000737:chemistry; D004789:Enzyme Activation; D006861:Hydrogen Peroxide / Q000737:chemistry / Q000378:metabolism; D006868:Hydrolysis; D007506:Iron-Sulfur Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism; D008297:Male; D008929:Mitochondria, Heart / Q000201:enzymology*; D016877:Oxidants / Q000737:chemistry; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D051381:Rats; D017207:Rats, Sprague-Dawley; D012402:Rotenone / Q000737:chemistry; D013481:Superoxides / Q000737:chemistry
chemical_list: D007506:Iron-Sulfur Proteins; D016877:Oxidants; D012402:Rotenone; D013481:Superoxides; D006861:Hydrogen Peroxide; D010450:Endopeptidases; D000154:Aconitate Hydratase