Photosystem II (PSII) is a huge membrane-protein complex consisting of 20 different subunits with a total molecular mass of 350 kDa for a monomer. It catalyses light-driven water oxidation at its catalytic centre, the oxygen-evolving complex (OEC)^1,2,3. The structure of PSII has been analysed at 1.9 Å resolution by synchrotron radiation X-rays, which revealed that the OEC is a Mn₄CaO₅ cluster organized in an asymmetric, ‘distorted-chair’ form⁴. This structure was further analysed with femtosecond X-ray free electron lasers (XFEL), providing the ‘radiation damage-free’⁵ structure. The mechanism of O=O bond formation, however, remains obscure owing to the lack of intermediate-state structures. Here we describe the structural changes in PSII induced by two-flash illumination at room temperature at a resolution of 2.35 Å using time-resolved serial femtosecond crystallography with an XFEL provided by the SPring-8 ångström compact free-electron laser. An isomorphous difference Fourier map between the two-flash and dark-adapted states revealed two areas of apparent changes: around the Q_B/non-haem iron and the Mn₄CaO₅ cluster. The changes around the Q_B/non-haem iron region reflected the electron and proton transfers induced by the two-flash illumination. In the region around the OEC, a water molecule located 3.5 Å from the Mn₄CaO₅ cluster disappeared from the map upon two-flash illumination. This reduced the distance between another water molecule and the oxygen atom O4, suggesting that proton transfer also occurred. Importantly, the two-flash-minus-dark isomorphous difference Fourier map showed an apparent positive peak around O5, a unique μ₄-oxo-bridge located in the quasi-centre of Mn1 and Mn4 (refs 4,5). This suggests the insertion of a new oxygen atom (O6) close to O5, providing an O=O distance of 1.5 Å between these two oxygen atoms. This provides a mechanism for the O=O bond formation consistent with that proposed previously^6,7.

Green to the core: Light from the Sun powers most life on today's Earth in some way. The core of the photosynthetic apparatus where charge separation occurs, the reaction center (RC), is thought to have originated a single time and diverged, yielding new kinds of complexes adapted to different tasks and environments. Chen et al. now present an important missing puzzle piece in our understanding of the evolution of RCs: a cryo–electron microscopy structure of the homodimeric type I RC from a green sulfur bacterium bound to a light-harvesting protein. The observed cofactor and pigment arrangement explain biochemical features of this RC and will aid in our understanding of how a single ancestral RC gave rise to the range of structures and functions seen in RCs today. Science , this issue p. eabb6350

Nitric oxide (NO) is an active and critical nitrogen oxide in the microbe-driven nitrogen biogeochemical cycle, and is of great interest to medicine and the biological sciences. As a gas molecule prior to oxygen, NO respiration represents an early form of energy generation via various reactions in prokaryotes. Major enzymes for endogenous NO formation known to date include two types of nitrite reductases in denitrification, hydroxylamine oxidoreductase in ammonia oxidation, and NO synthases (NOSs). While the former two play critical roles in shaping electron transport pathways in bacteria, NOSs are intracellular enzymes catalyzing metabolism of certain amino acids and have been extensively studied in mammals. NO interacts with numerous cellular targets, most of which are redox-active proteins. Doing so, NO plays harmful and beneficial roles by affecting diverse biological processes within bacterial physiology. Here, we discuss recent advances in the field, including NO-forming enzymes, the molecular mechanisms by which these enzymes function, physiological roles of bacterial NOSs, and regulation of NO homeostasis in bacteria.

Photosystem I (PSI) is a large protein supercomplex that catalyzes the light‐dependent oxidation of plastocyanin (or cytochrome c₆) and the reduction of ferredoxin. This catalytic reaction is realized by a transmembrane electron transfer chain consisting of primary electron donor (a special chlorophyll (Chl) pair) and electron acceptors A₀, A₁, and three Fe₄S₄ clusters, F_X, F_A, and F_B. Here we report the PSI structure from a Chl d‐dominated cyanobacterium Acaryochloris marina at 3.3 Å resolution obtained by single‐particle cryo‐electron microscopy. The A. marina PSI exists as a trimer with three identical monomers. Surprisingly, the structure reveals a unique composition of electron transfer chain in which the primary electron acceptor A₀ is composed of two pheophytin a rather than Chl a found in any other well‐known PSI structures. A novel subunit Psa27 is observed in the A. marina PSI structure. In addition, 77 Chls, 13 α‐carotenes, two phylloquinones, three Fe‐S clusters, two phosphatidyl glycerols, and one monogalactosyl‐diglyceride were identified in each PSI monomer. Our results provide a structural basis for deciphering the mechanism of photosynthesis in a PSI complex with Chl d as the dominating pigments and absorbing far‐red light.

Acaryochloris marina is a unique cyanobacterium using chlorophyll d (Chl d ) as its major pigment and thus can use far-red light for photosynthesis. Photosystem II (PSII) of A. marina associates with a number of prochlorophyte Chl-binding (Pcb) proteins to act as the light-harvesting system. We report here the cryo-electron microscopic structure of a PSII-Pcb megacomplex from A. marina at a 3.6-angstrom overall resolution and a 3.3-angstrom local resolution. The megacomplex is organized as a tetramer consisting of two PSII core dimers flanked by sixteen symmetrically related Pcb proteins, with a total molecular weight of 1.9 megadaltons. The structure reveals the detailed organization of PSII core consisting of 15 known protein subunits and an unknown subunit, the assembly of 4 Pcb antennas within each PSII monomer, and possible pathways of energy transfer within the megacomplex, providing deep insights into energy transfer and dissipation mechanisms within the PSII-Pcb megacomplex involved in far-red light utilization.

Anoxygenic phototrophic sulfur bacteria flourish in contemporary and ancient euxinic environments, driving the biogeochemical cycles of carbon and sulfur. However, it is unclear how these strict anaerobes meet their high demand for iron in iron-depleted environments. Here, we report that pyrite, a widespread and highly stable iron sulfide mineral in anoxic, low-temperature environments, can support the growth and metabolic activity of anoxygenic phototrophic sulfur bacteria by serving as the sole iron source under iron-depleted conditions. Transcriptomic and proteomic analyses revealed that pyrite addition substantially up-regulated genes and protein expression involved in photosynthesis, sulfur metabolism, and biosynthesis of organics. Anoxic microbial oxidation of pyritic sulfur and consequent destabilization of the pyrite structure were postulated to facilitate microbial iron acquisition. These findings advance our understanding of the survival strategies of anaerobes in iron-depleted environments and are important for revealing the previously underappreciated bioavailability of pyritic iron in anoxic environments and anoxic weathering of pyrite.