Intracellular potassium (K⁺) homeostasis is fundamental to cell viability. In addition to channels, K⁺ levels are maintained by various ion transporters. One major family is the proton-driven K⁺ efflux transporters, which in gram-negative bacteria is important for detoxification and in plants is critical for efficient photosynthesis and growth. Despite their importance, the structure and molecular basis for K⁺-selectivity is poorly understood. Here, we report ~3.1 Å resolution cryo-EM structures of the Escherichia coli glutathione (GSH)-gated K⁺ efflux transporter KefC in complex with AMP, AMP/GSH and an ion-binding variant. KefC forms a homodimer similar to the inward-facing conformation of Na⁺/H⁺ antiporter NapA. By structural assignment of a coordinated K⁺ ion, MD simulations, and SSM-based electrophysiology, we demonstrate how ion-binding in KefC is adapted for binding a dehydrated K⁺ ion. KefC harbors C-terminal regulator of K⁺ conductance (RCK) domains, as present in some bacterial K⁺-ion channels. The domain-swapped helices in the RCK domains bind AMP and GSH and they inhibit transport by directly interacting with the ion-transporter module. Taken together, we propose that KefC is activated by detachment of the RCK domains and that ion selectivity exploits the biophysical properties likewise adapted by K⁺-ion-channels.

Photosystem I (PSI) enables photo-electron transfer and regulates photosynthesis in the bioenergetic membranes of cyanobacteria and chloroplasts. Being a multi-subunit complex, its macromolecular organization affects the dynamics of photosynthetic membranes. Here we reveal a chloroplast PSI from the green alga Chlamydomonas reinhardtii that is organized as a homodimer, comprising 40 protein subunits with 118 transmembrane helices that provide scaffold for 568 pigments. Cryogenic electron microscopy identified that the absence of PsaH and Lhca2 gives rise to a head-to-head relative orientation of the PSI–light-harvesting complex I monomers in a way that is essentially different from the oligomer formation in cyanobacteria. The light-harvesting protein Lhca9 is the key element for mediating this dimerization. The interface between the monomers is lacking PsaH and thus partially overlaps with the surface area that would bind one of the light-harvesting complex II complexes in state transitions. We also define the most accurate available PSI–light-harvesting complex I model at 2.3 Å resolution, including a flexibly bound electron donor plastocyanin, and assign correct identities and orientations to all the pigments, as well as 621 water molecules that affect energy transfer pathways.

Plants, algae, and cyanobacteria convert light energy into chemical energy through the process of photosynthesis, fueling the planet and making life as we know it possible. Photosystem I (PSI) is one of the main photosynthetic complexes, responsible for this process. PSI uses the energy of light to transfer electrons from the soluble electron carrier plastocyanin, on the lumenal site of the thylakoid membrane, to ferrodoxin, on the stromal site of the membrane. Thus, playing a key role in the light dependent reactions. In order to survive many photosynthetic organisms need to be able to adapt to fluctuations in light and have adapted their photosynthetic machinery accordingly. In recent years many advances have been made in electron cryo-microscopy, making it possible to visualize many previously elusive photosynthetic complexes. This has brought a wealth of information on the structural adaptations of PSI.

In plants and algae, PSI is hosted by the chloroplast, a specialized organelle that houses the photosynthetic reactions. In the chloroplast, key components of PSI are synthesized by the chloroplasts own translation machinery: the chloroplast ribosome. Translation in the chloroplast is remarkable as it has to synchronize translation in two different genetic compartments as well as adapt to fluctuations in light. A glimpse of how this machinery has evolved to be able to fulfill all of these duties can be obtained from its three dimensional structure and its chloroplast specific features. However, despite all this structural information providing valuable clues as to the functioning of these systems, there are still many aspects of how they play a role that still remain unknown.

Photosystem I (PSI) is able to form different oligomeric states across various species. To reveal the structural basis for PSI dimerization and tetramerization, we structurally investigated PSI from the cyanobacterium Anabaena. This revealed a disrupted trimerization domain due to lack of the terminal residues of PsaL in the lumen, which resulted in PSI dimers with loose connections between monomers and weaker energy-coupled chlorophylls than in the trimer. At the dimer surface, specific phospholipids, cofactors and interactions in combination facilitated recruitment of another dimer to form a tetramer. Taken together, the relaxed luminal connections and lipid specificity at the dimer interface account for membrane curvature. PSI tetramer assembly appears to increase the surface area of the thylakoid membrane, which would contribute to PSI crowding. Photosystem I from the cyanobacterium Anabaena has a disrupted trimerization domain resulting in dimers with loose connections between monomers. Phospholipids and cofactors at the dimer surface facilitate further dimerization to form a tetramer.

Solar energy harnessed by oxygenic photosynthesis supports most of the life forms on Earth. In eukaryotes, photosynthesis occurs in chloroplasts and is achieved by membrane-embedded macromolecular complexes that contain core and peripheral antennae with multiple pigments. The structure of photosystem I (PSI) comprises the core and light-harvesting (LHCI) complexes, which together form PSI-LHCI. Here we determined the structure of PSI-LHCI from the salt-tolerant green alga Dunaliella salina using X-ray crystallography and electron cryo-microscopy. Our results reveal a previously undescribed configuration of the PSI core. It is composed of only 7 subunits, compared with 14-16 subunits in plants and the alga Chlamydomonas reinhardtii, and forms the smallest known PSI. The LHCI is poorly conserved at the sequence level and binds to pigments that form new energy pathways, and the interactions between the individual Lhca1-4 proteins are weakened. Overall, the data indicate the PSI of D. salina represents a different type of the molecular organization that provides important information for reconstructing the plasticity and evolution of PSI. The photosystem I light-harvesting complex from the salt-tolerant green alga Dunaliella salina has a core configuration composed of only seven subunits. This unusual molecular organization could inform the reconstruction of photosystem evolution.

Oxygenic photosynthesis produces oxygen and builds a variety of organic compounds, changing the chemistry of the air, the sea and fuelling the food chain on our planet. The photochemical reactions underpinning this process in plants take place in the chloroplast. Chloroplasts evolved ~1.2 billion years ago from an engulfed primordial diazotrophic cyanobacterium, and chlororibosomes are responsible for synthesis of the core proteins driving photochemical reactions. Chlororibosomal activity is spatiotemporally coupled to the synthesis and incorporation of functionally essential co-factors, implying the presence of chloroplast-specific regulatory mechanisms and structural adaptation of the chlororibosome^1,2. Despite recent structural information^3,4,5,6, some of these aspects remained elusive. To provide new insights into the structural specialities and evolution, we report a comprehensive analysis of the 2.9–3.1 Å resolution electron cryo-microscopy structure of the spinach chlororibosome in complex with its recycling factor and hibernation-promoting factor. The model reveals a prominent channel extending from the exit tunnel to the chlororibosome exterior, structural re-arrangements that lead to increased surface area for translocon binding, and experimental evidence for parallel and convergent evolution of chloro- and mitoribosomes.