Jonathan L. Cape, Michael K. Bowman, and David M. Kramer.
A semiquinone intermediate generated at the Qo site of the cytochrome bc1 complex: Importance for the Q-cycle and superoxide production.
PNAS. published 30 April 2007, 10.1073/pnas.0702621104
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2 comments:
I think this is a very important contribution, but it is unlikely that this semiquinone can be on the normal reaction pathway. More likely the semiquinone observed here is the first intermediate in the abnormal pathway leading to ROS production. Maybe the first step after the common intermediate (of the normal pathway and ROS production)invoked by Forquer et al. If this is the case, observation of this semiquinone does not exclude mechanisms involving a concerted reaction or a rapid quasi-equilibrium with a highly unstable semiquinone. In fact if it is on an aberrant pathway it cannot tell us much at all about the normal reaction.
The main importance may be as a tool for investigating the Qo site by advanced pulsed epr techniques, and what it can tell us about the mechanism of ROS production by Complex III.
The reason for doubting this semiquinone is part of the normal reaction is the oxygen sensitivity- If O2 greatly diminishes the amount of semiquinone, it means the reaction with O2 is much faster than the rate of production of the semiquinone, hence the former can't be rate-limiting in the pathway to superoxide: the rate-limiting step is earlier, perhaps the formation of the semiquinone.
This means that antimycin would not inhibit oxidation of ubiquinone in the presence of oxygen, nor inhibit cyt c reduction if electrons on ROS go quantitatively to cyt c.
That is the "O2-dependent, SOD-sensitive antimycin leak" would be ~100% rather than 2-5%.
If this semiquinone is the first step after the common intermediate, what is the common intermediate? Since this semiquinone shows no evidence of N ligation, maybe the common intermediate is a semiquinone still complexed with the Rieske which in a small fraction of cycles gets loose, uncoupling the spin-pair with Rieske or prolonging its life to make it detectable, and allowing it to react with oxygen.
Alternatively the common intermediate is not a semiquinone at all but the ubiquinol/oxidized Rieske "substrate complex" which is about to undergo the concerted reaction but, in a few percent of cycles, has an aberrant reaction reducing Rieske only, or a back-transfer of the electron from heme b(L), resulting in semiquinone.
-ed berry
Another way of looking at this problem- if the O2-sensitive SQ seen here is on the pathway of normal turnover, then normal uninhibited turnover could never be faster than the rate of superoxide production in the presence of antimycin. This can be seen by the following thought experiment:
Start with the OIR conditions of the experiment: reduced Q-pool, oxidized high-potential chain, low potential chain blocked by antimycin, and O2-saturated medium.
Formation of the semiquinone is going at its maximal rate, due to driving force from reduced Q and oxidized Rieske, yet it is not fast enough to keep up with the O2 reaction, so no semiquinone accumulates; there is no reversal of the semiquinone formation step.
Now remove the antimycin so cyt b competes with oxygen to accept the electron from the semiquinone. Say the reaction with cyt b is much faster, so superoxide production essentially stops. We can even remove the oxygen at this stage to ensure all the electrons go by the normal pathway. But since semiquinone formation was already going at its maximal rate, and was rate-limiting for superoxide production, it will still be rate limiting and limit the rate of turnover to that same rate.
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