Kinetics of charge recombination and distribution of the ubiquinone pool in reaction center - Light harvesting complexes purified from Rhodobacter sphaeroides

We have recently shown that in the reaction center – light harvesting complex 1 (RC-LH1) purified from the photosynthetic bacterium Rhodobacter sphaeroides, at pH=7.8, flash-induced P+QB- recombines with an average rate constant ( <k> = 0.3 s-1), significantly smaller than that measured in RC deprived of the LH1 ( <k> = 1 s-1). Since the kinetics of P+QA- recombination is unaffected by the presence of the antenna, the P+QB- state appears to be energetically stabilized in core complexes (1). The pH dependence of the P+QB- recombination kinetics has been analyzed in dimeric and monomeric forms of RC-LH1 and compared with that observed in RC deprived of the antenna. At 6.5 < pH < 8.5 recombination is essentially pH independent and significantly slower in all forms of core complexes as compared to LH1-deprived RCs. At increasing pH values, however, where the recombination rate increases, this stabilization effect decreases progressively and vanishes at pH > 10.5, indicating the involvement of protonatable groups. The recombination kinetics, moreover, becomes progressively distributed at pH > 9. The width of the rate constant distribution (sigma = 0.3 s-1 at pH < 9.0) increases by more than one order of magnitude at pH 11.0, suggesting a variety of conformations, possibly differing in the protonation state. The observed pH dependence of sigma could be explained when assuming that such conformations interconvert at a rate which is comparable to the rate of charge recombination at physiological pH values but is considerably lower at high pH values. Under this condition the conformational heterogeneity becomes therefore observable. A similar behaviour was observed in chromatophores of Rhodobacter capsulatus FJ2, a c2- and cyminus strain, in which the kinetics of P+QB- recombination could be accurately studied by avoiding any interference due to exogenous electron donors/acceptors. The lipid complement of the examined RC-LH1 complexes, determined by Inductively Coupled Plasma Emission Spectroscopy of phosphorous, ranges between 200 and 400 phospolipid molecules per RC. A large ubiquinone (UQ) pool, varying from 15 to 30 UQ molecules per RC was systematically found to be associated with the core complexes. When similar determinations are performed in chromatophores, it appears that the effective UQ concentration in the lipid phase of core complexes is at least three times higher than the average UQ concentration in the intact membrane. This finding argues strongly in favour of an in vivo heterogeneity in the distribution of the quinone pool within the chromatophore bilayer.