Iron-sulfur components of succinate dehydrogenase: stoichiometry and kinetic behavior in activated preparations.
Extensively or completely activated preparations of beef heart succinate dehydrogenase have been investigated by electron paramagnetic resonance (EPR) techniques at 6 to 97 K. Reductive titrations with dithionite and rapid kinetic studies were performed with various types of soluble and membrane-bound preparations of the enzyme. The following components were detected and their behavior analyzed: a free radical, presumably arising from the covalently bound flavin on reduction, two iron-sulfur centers of the ferredoxin type, the signals of which appear on reduction, and a highpotential iron-sulfur component, detectable in the oxidized state. The high-potential component was only detected in complex II and inner-membrane preparations. This component and one of the ferredoxin-type centers were present in amounts close to stoichiometric with the flavin and were reduced by substrate. The other ferredoxin-type center was present in amounts between 0.1 and 0.5 times that of the flavin and was reduced only by dithionite. Of the components reduced by succinate, however, only a fraction (up to 50% of the high-potential iron-sulfur center and 40-60% of the ferredoxin-type iron-sulfur center) was reduced within the turnover time of the enzymes; In complex II not more than about 10% of the flavin appeared in the semiquinone form at any time. Soluble, purified preparations behaved similarly except that the high-potential component was nearly or completely absent and extensive accumulation of the free radical occurred (up to 70 to 80% of the flavin) in titration and kinetic experiments. No significant difference was observed between the rates of semiquinone formation and the reduction of the ferredoxin-type or high-potential centers by the substrate. Also no qualitative differences in the properties studied in this work became apparent between prepatations containing 4 or 8 iron atoms, respectively.
D000818:Animals; D002417:Cattle; D004227:Dithionite; D004578:Electron Spin Resonance Spectroscopy; D004789:Enzyme Activation; D005288:Ferredoxins / Q000032:analysis; D005415:Flavins; D005609:Free Radicals; D007501:Iron / Q000032:analysis*; D007700:Kinetics; D008931:Mitochondria, Muscle / Q000201:enzymology*; D009206:Myocardium / Q000201:enzymology* / Q000648:ultrastructure; D010084:Oxidation-Reduction; D013385:Succinate Dehydrogenase / Q000032:analysis; D013455:Sulfur / Q000032:analysis*
D005288:Ferredoxins; D005415:Flavins; D005609:Free Radicals; D004227:Dithionite; D013455:Sulfur; D007501:Iron; D013385:Succinate Dehydrogenase