Current perspectives on proton and electron transfer pathways in photosystem II water oxidation

Photosystem II catalyzes the light-driven oxidation of water, progressing via sequential oxidation states (S-states) of the Mn4CaO5 cluster. Among structural snapshots of intermediate S-states obtained using X-ray free-electron laser (XFEL) crystallography, two-flash XFEL structures assigned to the S3 state reveal an additional oxygen atom (O6) near the O5 site of the cluster, leading to proposals that O6 is incorporated as a new substrate water molecule during the S2 to S3 transition. However, recent re-analyses of the XFEL data highlight potential complications, including conformational heterogeneity, refinement bias, and possible radiation-induced artifacts. In addition, many proposals have been put forwarded without evaluating associated proton and electron transfer processes, despite the fact that water oxidation involves the stepwise removal of protons and electrons. Here, we shed light on electron and proton transfer events during the photocycle by summarizing mechanistic proposals, including those in which O6 is not incorporated. If the remaining reduced site, Mn1(III), is oxidized during the S2 to S3 transition, this step encounters difficulties due to its high redox potential and poor electronic coupling with the electron acceptor, D1-Tyr161 (TyrZ). Efficient proton transfer requires pre-existing H-bond networks, which are absent near O5 and O6, imposing kinetic penalties on proton release. Assigning O6 as a substrate oxygen would imply that O5 is the other substrate, requiring its deprotonation earlier in the Kok cycle.