Infrared Determination of the Protonation State of a Key Histidine Residue in the Photosynthetic Water Oxidizing Center.

Photosynthetic water oxidation is performed at the Mn4CaO5 cluster in photosystem II (PSII). The protonation structures of amino acid residues and water molecules around the Mn4CaO5 cluster are crucial in water oxidation reactions. In this study, we determined the protonation state of a key His residue in water oxidation, D1-H337, that is directly hydrogen-bonded with the oxygen atom of the Mn4CaO5 cluster, using polarized attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy. Flash-induced polarized ATR-FTIR difference spectra upon the S1 → S2 transition of oriented PSII membranes showed broad negative and positive features at about 2600 and 2900 cm-1, respectively, with large dichroic ratios, accompanied by several minor peaks attributable to the Fermi resonance of a His NH vibration. Quantum mechanics/molecular mechanics (QM/MM) calculations well reproduced the characteristics of these features as the NτH stretching vibrations of D1-H337 in its protonated cation form. The spectral features were reversed in the S3 → S0 transition, indicating that this His remains protonated during the S-state cycle. The redox potential (Em) of the Mn4CaO5 cluster in the S1 → S2 transition, which was estimated from the QM/MM calculations, was found to be comparable to that of water oxidation when D1-H337 is protonated cation. It was thus concluded that the positive charge on the protonated D1-H337 plays an important role in retaining a high Em value of the Mn4CaO5 cluster throughout the reaction cycle to enable water oxidation.