Superoxide radical and iron modulate aconitase activity in mammalian cells.
Aconitase is a member of a family of iron-sulfur-containing (de)hydratases whose activities are modulated in bacteria by superoxide radical (O2-.)-mediated inactivation and iron-dependent reactivation. The inactivation-reactivation of aconitase(s) in cultured mammalian cells was explored since these reactions may impact important and diverse aconitase functions in the cytoplasm and mitochondria. Conditions which increase O2-. production including exposure to the redox-cycling agent phenazine methosulfate (PMS), inhibitors of mitochondrial ubiquinol-cytochrome c oxidoreductase, or hyperoxia inactivated aconitase in mammalian cells. Overproduction of mitochondrial Mn-superoxide dismutase protected aconitase from inactivation by PMS or inhibitors of ubiquinol-cytochrome c oxidoreductase, but not from normobaric hyperoxia. Aconitase activity was reactivated (t1/2 of 12 +/- 3 min) upon removal of PMS. The iron chelator deferoxamine impaired reactivation and increased net inactivation of aconitase by O2-.. The ability of ubiquinol-cytochrome c oxidoreductase-generated O2-. to inactivate aconitase in several cell types correlated with the fraction of the aconitase activity localized in mitochondria. Extracellular O2-. generated with xanthine oxidase did not affect aconitase activity nor did exogenous superoxide dismutase decrease aconitase inactivation by PMS. The results demonstrate a dynamic and cyclical O2-.-mediated inactivation and iron-dependent reactivation of the mammalian [4Fe-4S] aconitases under normal and stress conditions and provide further evidence for the membrane compartmentalization of O2-..
Gardner
P R
PR
Department of Pediatrics, National Jewish Center for Immunology and Respiratory Medicine, Denver, Colorado 80206, USA.
Raineri
I
I
Epstein
L B
LB
White
C W
CW
D000154:Aconitate Hydratase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D000968:Antimycin A / Q000494:pharmacology; D002451:Cell Compartmentation; D002460:Cell Line; D002462:Cell Membrane / Q000201:enzymology; D003593:Cytoplasm / Q000201:enzymology; D014450:Electron Transport Complex III / Q000037:antagonists & inhibitors; D004793:Enzyme Reactivators / Q000494:pharmacology; D005609:Free Radicals; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007501:Iron / Q000494:pharmacology*; D008773:Methylphenazonium Methosulfate / Q000494:pharmacology; D051379:Mice; D008928:Mitochondria / Q000201:enzymology; D010100:Oxygen / Q000494:pharmacology; D051381:Rats; D013482:Superoxide Dismutase / Q000378:metabolism; D013481:Superoxides / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D004793:Enzyme Reactivators; D005609:Free Radicals; D013481:Superoxides; D008773:Methylphenazonium Methosulfate; D000968:Antimycin A; D006861:Hydrogen Peroxide; D007501:Iron; D013482:Superoxide Dismutase; D000154:Aconitate Hydratase; D014450:Electron Transport Complex III; D010100:Oxygen