Investigation of the functional role of Ca²⁺ in the oxygen-evolving complex of photosystem II: A pH-dependence study of the substitution of Ca²⁺ by Sr²⁺

The oxygen-evolving complex (OEC) of photosystem II (PSII) is the catalytic site for the oxidation of water to O₂ in photosynthesis. The OEC consists of a tetranuclear manganese (Mn₄) cluster with calcium and chloride ions functioning as essential cofactors. Previous studies have shown that substitution of Ca²⁺ in the OEC by various divalent and trivalent metal ions results in the loss of oxygen-evolution activity of PSII. Sr²⁺ is the only ion that has been found to restore the oxygen-evolving activity in PSII. Although several models have been proposed for the mechanism of water oxidation in PSII, the functional role of Ca ²⁺ remains unclear. In order to test the proposal that Ca ²⁺ functions as a Lewis acid in water oxidation chemistry, the pH dependence of the oxygen-evolution activity of intact Ca²⁺-containing PSII and Sr²⁺-substituted PSII has been investigated. In both samples, the pH dependence exhibits a bell-shaped curve. Sr²⁺ substitution shifts the peak of the curve to higher pH in comparison to Ca ²⁺-containing PSII. The pH-dependent O₂-evolving rates of these two samples are fit to a diprotic model in which protonation of an essential basic group at acidic pH inhibits activity (pKa₁) and deprotonation of an essential acidic group at higher pH inhibits activity (pKa₂). Sr²⁺ substitution has no effect on pKa ₂. This result suggests that the high pH inhibition may be on the electron-acceptor side of PSII. On the other hand, Sr²⁺ substitution causes pKa₁ to be shifted to higher pH by 1.0 pH unit. This pH-shift indicates a direct role of Ca²⁺/Sr²⁺ in the OEC. The functional role of Ca²⁺ in oxygen evolution in PSII is discussed based on these results.