Photosynthetic water oxidation occurs at the oxygen-evolving complex (OEC) of Photosystem II (PSII). The OEC, which contains a Mn₄CaO₅ inorganic cluster ligated by oxides, waters and amino-acid residues, cycles through five redox intermediates known as S_i states (i = 0–4). The electronic and structural properties of the transient S₄ intermediate that forms the O–O bond are not well understood. In order to gain insight into how water is activated for O–O bond formation in the S₄ intermediate, we have performed a detailed analysis of S-state dependent substrate water binding kinetics taking into consideration data from Mn coordination complexes. This analysis supports a model in which the substrate waters are both bound as terminal ligands and react via a water-nucleophile attack mechanism.