Ubisemiquinone is the electron donor for superoxide formation by complex III of heart mitochondria

Abstract

Much evidence indicates that superoxide is generated from O 2 in a cyanide-sensitive reaction involving a reduced component of complex III of the mitochondrial respiratory chain, particularly when antimycin A is present. Although it is generally believed that ubisemiquinone is the electron donor to O 2, little experimental evidence supporting this view has been reported. Experiments with succinate as electron donor in the presence of antimycin A in intact rat heart mitochondria, which contain much superoxide dismutase but little catalase, showed that myxothiazol, which inhibits reduction of the Rieske iron-sulfur center, prevented formation of hydrogen peroxide, determined spectrophotometrically as the H 2O 2-peroxidase complex. Similarly, depletion of the mitochondria of their cytochrome c also inhibited formation of H 2O 2, which was restored by addition of cytochrome c. These observations indicate that factors preventing the formation of ubisemiquinone also prevent H 2O 2 formation. They also exclude ubiquinol, which remains reduced under these conditions, as the reductant of O 2. Since cytochrome b also remains fully reduced when myxothiazol is added to succinate and antimycin A-supplemented mitochondria, reduced cytochrome b may also be excluded as the reductant of O 2. These observations, which are consistent with the Q-cycle reactions, by exclusion of other possibilities leave ubisemiquinone as the only reduced electron carrier in complex III capable of reducing O 2 to O 2 −.