RAPID PROTON-TRANSFER UNDER FLASHING LIGHT AT BOTH FUNCTIONAL SIDES OF DARK-ADAPTED PHOTOSYSTEM-II PARTICLES

By exposing dark-adapted Photosystem II core particles to a series of light flashes, we aimed at kinetic resolution of proton release during the four steps of water oxidation. The signal-to-noise ratio was improved by averaging under repetitive dark adaptation. The previously observed kinetic damping of pH-transients by particle aggregation was prevented by detergent. The complicating superimposition of protolytic events at the donor side (water oxidation) and at the acceptor side (quinone oxide-reduction) was unravelled by characterizing the rate constants of electron and proton transfer at the acceptor side (Q(A)(-) . nH(+) + DCBQ --> Q(A) + DCBQ(-) + nH(+): k = 1.7 . 10(6) M(-1) s(-1)//2 DCBQ(-) + 2H(+) --> DCBQ + DCBQH(2): k = 4 . 10(8) M(-1) s(-1)). Contrasting with the pronounced period of four oscillations of the oxygen-evolving centre, the extent of proton release was practically constant. The apparent half-rise time of the stepped acidification was shortened upon lowering of the pH (250 mu s at pH 7.5, 70 mu s at pH 6.0 and 12 mu s at pH 5.2). This kinetic behaviour was independent of the nature and the concentration of the added pH-indicator. We conclude that this reflects the protolysis of several electrostatically interacting acids at the surface of the protein in response to a new positive charge on Y-Z(+), and persisting upon electron transfer from the manganese cluster to Y-Z(+).