Structural evidence for intermediates during O formation in photosystem II.

In natural photosynthesis, the light-driven splitting of water into electrons, protons and molecular oxygen forms the first step of the solar-to-chemical energy conversion process. The reaction takes place in photosystem II, where the MnCaO cluster first stores four oxidizing equivalents, the S to S intermediate states in the Kok cycle, sequentially generated by photochemical charge separations in the reaction center and then catalyzes the O-O bond formation chemistry. Here, we report room temperature snapshots by serial femtosecond X-ray crystallography to provide structural insights into the final reaction step of Kok's photosynthetic water oxidation cycle, the S→[S]→S transition where O is formed and Kok's water oxidation clock is reset. Our data reveal a complex sequence of events, which occur over micro- to milliseconds, comprising changes at the MnCaO cluster, its ligands and water pathways as well as controlled proton release through the hydrogen-bonding network of the Cl1 channel. Importantly, the extra O atom O, which was introduced as a bridging ligand between Ca and Mn1 during the S→S transition, disappears or relocates in parallel with Y reduction starting at approximately 700 μs after the third flash. The onset of O evolution, as indicated by the shortening of the Mn1-Mn4 distance, occurs at around 1,200 μs, signifying the presence of a reduced intermediate, possibly a bound peroxide.