We focus on mechanisms that promote energy conserving electron and proton translocation through ubiquinol-cytochrome c oxidoreductase (bc1 complex). We shall make use of experimental systems that offer the means to activate the system by light flashes via a on a bacterial photochemical reaction center. Such systems can provide valuable information on a rapid timescale, of kinetics of oxidation/reduction and protonation/deprotonation of ratios of electron and proton movements and, in combination with redox potentiometry, of the fundamental properties of the active redox components. The first part is aimed at the mechanism of action of bc1 complexes in vivo and in vitro. The principal systems are: a) the native membranes from photosynthetic bacteria (Rhodopseudomonas sphaeroides and capsulata); b) hybrid constructions in vitro of well-characterized isolated reaction centers (Rps. sphaeroides), cytochrome c and bc1 complexes, (beef heart mitochondria). The in vivo photosynthetic bacterial membranes provide a coupled photon-electron-proton-phosphorylation membrane system that possesses the usual manipulative opportunities offered by bacteria, including growth/media manipulations and mutants blocked in the bc1, complex. The in vitro hybrid constructions provide wide experimental flexibility and provide the means of achieving more ordered, densely packed mono- and multilayer arrays of the hybrid systems on glass or electrodes, for structurally related spectrometric analysis and direct measurement and control of charge movements across the monolayer profile. The second part approaches the bc1 complex from a different direction: from the study of a key component--ubiquinone. Quinonoid compounds will be studied from the standpoint of their structure, their substituents, electrochemistry, and hydrophobic properties. Quinonoid compounds will be used to probe the bc1 complex catalytic sites of ubiquinone oxidation-reduction to determine the parameters that govern the association of molecule and reaction site(s). Further efforts will be made to develop a family of bc complexes, systematically altered in the electrochemistry of the replacement quinones. This approach should strongly augment features of the first theme of the proposal. Moreover, it will provide the basis for a much needed, alternate approach to obtaining kinetic and thermodynamic information from the systematically altering free energy gaps between reactants and add another dimension to the study of electric field effects on the bc1 complexes oriented as mono- and multilayers between electrode surfaces.
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