Rapid redox equilibrium between the mitochondrial Q pool and cytochrome b during triphasic reduction of cytochrome b by succinate.
The reliability of monitoring the redox reactions of cytochrome b using the different wavelengths employed by different authors has been reexamined. It was found that 562-575 nm is suitable in succinate: cytochrome c reductase but not in mitochondria, in which case 562-540 nm is a better pair. Direct optical measurements of the redox reaction kinetics of the mitochondrial Q pool using a commercial dual-wavelength spectrophotometer are possible when succinate is used as the electron donor. Using the correct wavelength pair, and with malonate to slow down the electron input, the reduction course of cytochrome b was still triphasic but a plateau or a turn replaced the oxidation phase previously reported by several authors. At the same time, the reduction course of the Q pool was also triphasic, and in perfect match with that of cytochrome b. Destruction of the Rieske iron-sulfur cluster by British anti-Lewisite (BAL) + O2 treatment or prereduction of the high-potential components made the reduction of both Q and b monophasic. The plot of log (Q/QH2) against log (b3+/b2+) gave a straight line with an n value of 1.7 for cytochrome b at pH 7.4. This n value rose to 2.0 at pH 6.5 and dropped to 1.4 at pH 8.5. On the other hand, the mid-point potential of cytochrome b relative to that of the Q pool remained essentially unchanged between pH 6.5 and 8.4. BAL treatment had a small effect on the midpoint potential of cytochrome b relative to that of the Q pool and had no effect on the n value. Addition of quinone homologues and analogues extended the plateau phase in the reduction of cytochrome b, but exogenous quinones did not equilibrate rapidly with cytochrome b. It was concluded that the appearance of the plateau between the two reduction phases of Q and b is caused by the rapid delivery of electrons to the high-potential components of the respiratory chain as envisaged in the Q cycle; the unexpected n value for cytochrome b suggests a concerted reduction by QH2 of two species of cytochromes b-562.
D000818:Animals; D000968:Antimycin A / Q000031:analogs & derivatives / Q000494:pharmacology; D003573:Cytochrome b Group / Q000378:metabolism*; D029968:Escherichia coli Proteins; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008928:Mitochondria / Q000378:metabolism*; D010084:Oxidation-Reduction; D011809:Quinones / Q000494:pharmacology; D013053:Spectrophotometry; D013384:Succinate Cytochrome c Oxidoreductase / Q000378:metabolism; D013386:Succinates / Q000494:pharmacology*; D019802:Succinic Acid; D013552:Swine; D014451:Ubiquinone / Q000378:metabolism*
D003573:Cytochrome b Group; D029968:Escherichia coli Proteins; D011809:Quinones; D013386:Succinates; C032456:antimycin; D014451:Ubiquinone; D000968:Antimycin A; C017850:cytochrome b562, E coli; D019802:Succinic Acid; D013384:Succinate Cytochrome c Oxidoreductase