Proton Release Process during the S₂‑to‑S₃ Transition of Photosynthetic Water Oxidation As Revealed by the pH Dependence of Kinetics Monitored by Time-Resolved Infrared Spectroscopy

Photosynthetic water oxidation takes place at the Mn₄CaO₅ cluster in photosystem II via a light-driven cycle of intermediates called S states (S₀–S₄). Clarifying how electron and proton transfer reactions are coupled with each other in the S₂ → S₃ transition, which occurs just before O–O bond formation, is crucial for understanding the water oxidation mechanism. Here, we investigated the pH dependence of the kinetics of the S₂ → S₃ transition using time-resolved infrared (TRIR) spectroscopy to identify the proton release phase in this transition. TRIR measurements of Y_D-less PSII core complexes from the D2-Y160F mutant of Thermosynechococcus elongatus showed that the last phase in this transition (τ ∼ 350 μs at pH 6) was strongly dependent on pH, and its time constant at pH 5 was larger than that at pH 8 by a factor of >3. In contrast, the earlier phase with a time constant of ∼100 μs was virtually independent of pH. These results strongly support the view that proton release is a rate-limiting step of the proton-coupled electron transfer in the last phase of the S₂ → S₃ transition. This proton release enables electron transfer by removing an excessive positive charge from the catalytic center and hence decreasing its redox potential.