Exploring the interdependence of calcium and chloride activation of O₂ evolution in photosystem II

Calcium and chloride are activators of oxygen evolution in photosystem II (PSII), the light-absorbing water oxidase of higher plants, algae, and cyanobacteria. Calcium is an essential part of the catalytic Mn₄CaO₅ cluster that carries out water oxidation and chloride has two nearby binding sites, one of which is associated with a major water channel. The co-activation of oxygen evolution by the two ions is examined in higher plant PSII lacking the extrinsic PsbP and PsbQ subunits using a bisubstrate enzyme kinetics approach. Analysis of three different preparations at pH 6.3 indicates that the Michaelis constant, K_M, for each ion is less than the dissociation constant, K_S, and that the affinity of PSII for Ca²⁺ is about ten-fold greater than for Cl^-, in agreement with previous studies. Results are consistent with a sequential binding model in which either ion can bind first and each promotes the activation by the second ion. At pH 5.5, similar results are found, except with a higher affinity for Cl^- and lower affinity for Ca²⁺. Observation of the slow-decaying Tyr Z radical, Y_Z•, at 77 K and the coupled S₂Y_Z• radical at 10 K, which are both associated with Ca²⁺ depletion, shows that Cl^- is necessary for their observation. Given the order of electron and proton transfer events, this indicates that chloride is required to reach the S₃ state preceding Ca²⁺ loss and possibly for stabilization of Y_Z• after it forms. Interdependence through hydrogen bonding is considered in the context of the water environment that intervenes between Cl^- at the Cl-1 site and the Ca²⁺/Tyr Z region.