The slow rise of the flash-light-induced alkalization by photosystem II of the suspending medium of thylakoids is reversibly related to thylakoid stacking.

We studied the kinetics of flash-induced proton uptake at the reducing site of Photosystem II with Cresol red as indicator for pH transients in the suspending medium. The rise of the alkalization which was observed when Photosystem II was hidden in the stacked regions of the thylakoid membranes was much slower (100 ms) than the reduction of the bound quinones (less than 1 ms). We asked for the delay mechanism. We found that the rise of the alkalization became biphasic if thylakoids were unstacked. This was reversed upon restacking: The portion of the fast phase (half-rise time, 2.7 ms) increased, if the concentration of Mg high 2 plus was lowered. After EDTA-treatment we observed solely fast proton uptake. Experiments with dark-adapted chloroplasts showed that the biphasicity was not attributable to the alternating transitions of the bound quinone acceptors through the semiquinone and the hydroquinone stages. The dependence of fast proton uptake on the degree of membrane stacking was o n line with our previous proposal (Hong and Junge (1983) Biochim.Biophys. Acta 722, 197-205) that the propagation of a pH pulse in the narrow gaps between stacked membranes was slowed down by multiple reactions of protons with fixed buffering groups. This concept was corroborated by a theory which was given in the subsequent article.