The impact of membrane-targeted cations on energy conversion
in membrane vesicles from Rb. capsulatus

M.A. Kozloval, A.Y. Mulkidjanian2,3
1Institute of Mitoengineering, Moscow State University, Moscow 119899, Russia
2Division of Biophysics, Faculty of Biology/Chemistry, University of Osnabrück, D-49069 Osnabrück, Germany
3A.N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow 119899, Russia.

We have studied the impact of mitochondrially targeted penetrating cations on energy conversion in membrane vesicles from phototrophic bacteria Rb. capsulatus (chromatophores). The energy-converting enzymes of purple bacteria are closely related to their mitochondrial counterparts, but can be synchronized by short flashes of light. The same membranes contain electrochromic pigments, mostly carotenoids, which can serve as internal voltmeters.
The mitochondrially targeted cations, did not affect notably the flash-triggered generation of membrane potential (at least when added at < 1 μM), but accelerated its decay. The similar uncoupling efficiency of SkQ1, MitoQ and C12TPP indicates the involvement of the TPP moiety in the proton translocation. This uncoupling action could be partly prevented by depletion of chromatophore preparation from fatty acids (via pre-washing with bovine serum albumin, BSA). Addition of palmitic acid to the BSA-treated chromatophores partially reversed the effect of BSA. We consider these data as an evidence for the transmembrane, H+-conducting fatty acid cycling mediated by penetrating cations, as suggested by Skulachev and co-workers (Severin et al. 2010).
Neither SkQ nor MitoQ affected the flash induced redox-changes of cytochrome b in the cytochrome bc1 complex. Accordingly, these cations which, most likely, can interact only with the quinone-reducing center N of the complex (Gu et al., 2000), did not block the electron flow out of this center.
When added in concentration of > 5μM over antimycin A, SkQ1 could serve as an electron shuttle, overcoming the antimycin block. This observation explains why the addition of SkQ1 over antimycin induces a ROS burst in mitochondria.
The studies of the Zn-treated chromatophores, where the oxidation of cytochrome b was retarded (Skulachev et al., 1967, Klishin et al. 2002, Mulkidjanian, 2007) have shown that MitoQ and SkQ, when addded at > 1 μM slightly slowed down the reduction of heme bh. Any slowing of this reaction, however, should be accompanied by an increase in the ROS production.
We suggest that at high concentrations MitoQ and SkQ can bind to the cytochrome bc1 complex and affect the mechanism of ubiquinol oxidation in center P. This effect could account for the pro-oxidant action of the mitochondrially targeted cations.

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Homo Sapiens Liberatus Workshop, Moscow State University, May 2010