The respiratory chain of plant mitochondria XIII. Redox state changes of cytochrome b562 in mung bean seedling mitochondria treated with antimycin A

Abstract

1. 1. The response to antimycin A of the redox state changes of cytochrome b 562, the cytochrome b in mung bean seedling mitochondria with long wavelength absorbance maximum and low midpoint potential of −77 mV independent of mitochondrial energy state, has been examined with regard to possible spectral shifts, with regard to the energy state of the mitchondria, and with regard to the redox state of the cytochromes c. 2. 2. Absorbance changes observed at 566 to 540 nm on addition of antimycin A to the mitochondrial suspension correspond primarily to changes in the redox state of cytochrome b 562. Spectral shifts contribute little or nothing to these changes. 3. 3. When a suspension of antimycin A-treated mung bean mitochondria becomes anaerobic with succinate as substrate through the alternate oxidase pathway characteristic of plant mitochondria, cytochrome b 562, which is highly reduced in the aerobic steady state, becomes partially oxidized. The degree of reoxidation is maximal in the presence of ATP and decreased by the presence of uncoupler or oligomycin. Reoxidation of the anaerobic mitochondrial suspension causes re-reduction of cytochrome b 562. 4. 4. The oxidation of cytochrome b 562 in antimycin A-treated mitochondria in the presence of ATP on transition to anaerobiosis is synchronous with the reduction of cytochrome c 549 (corresponding to c 1 in mammalian mitochondria) observed during this transition. Reduction of highly oxidized cytochrome c 549 in antimycin A-treated mitochondria by addition of sulfide during the aerobic steady state results in a corresponding oxidation of highly reduced cytochrome b 562. 5. 5. Oxidation of the b cytochromes by means of the alternate oxidase, which operates on the substrate side of these cytochromes with reference to cytochrome oxidase, is very slow in the presence of antimycin A when the c cytochromes are oxidized. 6. 6. It is concluded that b 562 and antimycin A form a stabilized complex analogous to the cytochrome a 3/CO complex, in which the reduced form is energetically favored when the c cytochromes are highly oxidized. The midpoint potential of the stabilized complex would be more positive than that of uncomplexed cytochrome b 562 by an amount determined by the free energies of formation of the antimycin A complex with oxidized and reduced b 562, as is the case with the CO complex of cytochrome a 3. When the c cytochromes become reduced, the stabilized cytochrome b 562/antimycin A complex relaxes, and in this form resumes a more negative midpoint potential. The cytochrome then is reoxidized by equilibration with oxidized carriers on its substrate side. It is proposed that this mechanism is also applicable to the changes in redox state of cytochrome b T observed in solubilized mitochondria preparations from heart muscle treated with antimycin A.