Photosynthetic reaction centers produce and export oxidizing and reducing equivalents in expense of absorbed light energy. The formation of fully reduced quinone (quinol) requires a strict (1:1) stoichiometric ratio between the electrons and H+ ions entering the protein. The steady-state rates of both transports were measured separately under continuous illumination in the reaction center from the photosynthetic bacterium Rhodobacter sphaeroides. The uptake of the first proton was retarded by different methods and made the rate-limiting reaction in the photocycle. As expected, the rate constant of the observed proton binding remained constant (7 s-1), but that of the cytochrome photooxidation did show a remarkably large increase from 14 to 136 s-1 upon increase of the exciting light intensity up to 5 W/cm 2 (808 nm) at pH 8.4 in the presence of NiCl2. This corresponds to about 20:1 (e-:H+) stoichiometric ratio. The observed enhancement is linearly proportional to the light intensity and the rate constant of the proton uptake by the acceptor complex and shows saturation character with quinone availability. For interpretation of the acceleration of cytochrome turnover, an extended model of the photocycle is proposed. A fraction of photochemically trapped RC can undergo fast (> 103 s -1) conformational change where the semiquinone loses its high binding affinity (the dissociation constant increases by more than 5 orders of magnitude) and dissociates from the QB binding site of the protein with a high rate of 4000 s-1. Concomitantly, superoxide is being produced. No H+ ion is taken up, and no quinol is created by the photocycle which is operating in about 25% of the reaction centers at the highest light intensity (5500 s-1) and slowest proton uptake (3.5 s-1) used in our experiments. The possible physical background of the light-induced conformational change and the relationship between the energies of dissociation and redox changes of the quinone in the QB binding sites are discussed.
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