Superoxides from mitochondrial complex III: The role of manganese superoxide dismutase

In this report we show that ubiquinone cytochrome c reductase (complex III) from isolated rat heart mitochondria when inhibited with antimycin A, prod uces a large amount of superoxide as measured by the chemiluminescent probe coelenterazine. When mitochondria are inhibited with myxothiazol or stigma tellin, there is no detectable formation of superoxide. The antimycin A-sen sitive free radical production can be dramatically reduced using either myx othiazol or stigmatellin. This suggests that the antimycin A-sensitive gene ration of superoxides originates primarily from the Q(o) semiubiquinone. Wh en manganese superoxide dismutase depleted submitochondrial particles (SMP) were inhibited with myxothiazol or stigmatellin, a large superoxide signal was observed. These two inhibitors likely increase the concentration of th e Q(i) semiquinone at the N center. The antimycin A-sensitive signal can, i n the case of both the mitochondria and the SMP, be dissipated by the addit ion of copper zinc superoxide dismutase, suggesting that the measured coele nterazine signal was a result of superoxide production. Taken together, thi s data suggests that free radicals generated from the Q(i) species are more effectively eliminated by MnSOD in intact mitochondria. (C) 2000 Elsevier Science Inc.