Location of chloride and its possible functions in oxygen-evolving photosystem II revealed by X-ray crystallography

The chloride ion, Cl^-, is an essential cofactor for oxygen evolution of photosystem II (PSII) and is closely associated with the Mn ₄Ca cluster. Its detailed location and function have not been identified, however. We substituted Cl^- with a bromide ion (Br ^-) or an iodide ion (I^-) in PSII and analyzed the crystal structures of PSII with Br^- and I^- substitutions. Substitution of Cl^- with Br^- did not inhibit oxygen evolution, whereas substitution of Cl^- with I^- completely inhibited oxygen evolution, indicating the efficient replacement of Cl ^- by I^-. PSII with Br^- and I^- substitutions were crystallized, and their structures were analyzed. The results showed that there are 2 anion-binding sites in each PSII monomer; they are located on 2 sides of the Mn₄Ca cluster at equal distances from the metal cluster. Anion-binding site 1 is close to the main chain of D1-Glu-333, and site 2 is close to the main chain of CP43-Glu-354; these 2 residues are coordinated directly with the Mn₄Ca cluster. In addition, site 1 is located in the entrance of a proton exit channel. These results indicate that these 2 Cl^- anions are required to maintain the coordination structure of the Mn₄Ca cluster as well as the proposed proton channel, thereby keeping the oxygen-evolving complex fully active.