Probing the proton release by Photosystem II in the S₁ to S₂ high-spin transition

The stoichiometry and kinetics of the proton release were investigated during each transition of the S-state cycle in Photosystem II (PSII) from Thermosynechococcus elongatus containing either a Mn₄CaO₅ (PSII/Ca) or a Mn₄SrO₅ (PSII/Sr) cluster. The measurements were done at pH 6.0 and pH 7.0 knowing that, in PSII/Ca at pH 6.0 and pH 7.0 and in PSII/Sr at pH 6.0, the flash-induced S₂-state is in a low-spin configuration (S₂^LS) whereas in PSII/Sr at pH 7.0, the S₂-state is in a high-spin configuration (S₂^HS) in half of the centers. Two measurements were done; the time-resolved flash dependent i) absorption of either bromocresol purple at pH 6.0 or neutral red at pH 7.0 and ii) electrochromism in the Soret band of P_D1 at 440 nm. The fittings of the oscillations with a period of four indicate that one proton is released in the S₁ to S₂^HS transition in PSII/Sr at pH 7.0. It has previously been suggested that the proton released in the S₂^LS to S₃ transition would be released in a S₂^LSTyr_Z^• → S₂^HSTyr_Z^• transition before the electron transfer from the cluster to Tyr_Z^• occurs. The release of a proton in the S₁Tyr_Z^• → S₂^HSTyr_Z transition would logically imply that this proton release is missing in the S₂^HSTyr_Z^• to S₃Tyr_Z transition. Instead, the proton release in the S₁ to S₂^HS transition in PSII/Sr at pH 7.0 was mainly done at the expense of the proton release in the S₃ to S₀ and S₀ to S₁ transitions. However, at pH 7.0, the electrochromism of P_D1 seems larger in PSII/Sr when compared to PSII/Ca in the S₃ state. This points to the complex link between proton movements in and immediately around the Mn₄ cluster and the mechanism leading to the release of protons into the bulk.