Methyl-Coenzyme M Reductase (MCR) catalyzes the biosynthesis of methane in methanogenic archaea, using a catalytic Ni-centered Cofactor F430 in its active site. It also catalyzes the reverse reaction, that is, the anaerobic activation and oxidation, including the cleavage of the CH bond in methane. Because methanogenesis is the major source of methane on earth, understanding the reaction mechanism of this enzyme can have massive implications in global energy balances. While recent publications have proposed a radical-based catalytic mechanism as well as novel sulfonate-based binding modes of MCR for its native substrates, the structure of the active state of MCR, as well as a complete characterization of the reaction, remain elusive. Previous attempts to structurally characterize the active MCR-Ni(I) state have been unsuccessful due to oxidation of the redox- sensitive catalytic Ni center. Further, while many cryo structures of the inactive Ni(II)-enzyme in various substrates-bound forms have been published, no room temperature structures have been reported, and the structure and mechanism of MCR under physiologically relevant conditions is not known. In this study, we report the first room temperature structure of the MCRred1-silent Ni(II) form using an X-ray Free-Electron Laser (XFEL), with simultaneous X-ray Emission Spectroscopy (XES) and X-ray Diffraction (XRD) data collection. In celebration of the seminal contributions of inorganic chemist Dick Holm to our understanding of nickel-based catalysis, we are honored to announce our findings in this special issue dedicated to this remarkable pioneer of bioinorganic chemistry.

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 Mn₄CaO₅ cluster first stores four oxidizing equivalents, the S₀ to S₄ 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^1-3. 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 S₃→[S₄]→S₀ transition where O₂ 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 Mn₄CaO₅ 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_x, which was introduced as a bridging ligand between Ca and Mn1 during the S₂→S₃ transition^4-6, disappears or relocates in parallel with Y_z reduction starting at approximately 700 μs after the third flash. The onset of O₂ 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.

Photosystem I (PS I) has a symmetric structure with two highly similar branches of pigments at the center that are involved in electron transfer, but shows very different efficiency along the two branches. We have determined the structure of cyanobacterial PS I at room temperature (RT) using femtosecond X-ray pulses from an X-ray free electron laser (XFEL) that shows a clear expansion of the entire protein complex in the direction of the membrane plane, when compared to previous cryogenic structures. This trend was observed by complementary datasets taken at multiple XFEL beamlines. In the RT structure of PS I, we also observe conformational differences between the two branches in the reaction center around the secondary electron acceptors A_1A and A_1B. The π-stacked Phe residues are rotated with a more parallel orientation in the A-branch and an almost perpendicular confirmation in the B-branch, and the symmetry breaking PsaB-Trp673 is tilted and further away from A_1A. These changes increase the asymmetry between the branches and may provide insights into the preferential directionality of electron transfer.