Detection of structural changes upon S1-to-S2 transition in the oxygen-evolving manganese cluster in photosystem II by light-induced Fourier transform infrared difference spectroscopy.

The light-induced Fourier transform infrared (FT-IR) difference spectrum between the S1 and S2 states of the O2-evolving photosystem II (PSII) was obtained for the first time. Detection of an S2/S1 difference spectrum virtually free from contributions by the acceptor-side signals was achieved by employing an exogenous electron acceptor, potassium ferricyanide, to trypsin-treated PSII membranes and using one-flash excitation at 250 K. A synthetic difference spectrum obtained by adding this S2/S1 spectrum to the QA-/QA spectrum measured with Mn-depleted PSII was almost identical with the difference spectrum of the S2QA-/S1QA charge separation measured with untreated PSII. This successful simulation verifies the correctness of the S2/S1 spectrum thus obtained. The observed S2/S1 spectrum reflects the structural changes within the water-oxidizing Mn cluster upon the S1-to-S2 transition, most probably changes in vibrational modes of ligands coordinating to the Mn ion(s) that is (are) oxidized upon the S2 formation and/or changes in protein conformation. The present results demonstrate that FT-IR difference spectroscopy is a promising method to investigate the structure of the intermediates of the Mn cluster involved in photosynthetic water oxidation.