Biology subjects

Schuller, J.

Publications and source records attributed to Schuller, J..

4 recordsLinked to original sources

bioRxiv2025

Structural insights into late-stage photosystem II assembly by Psb32

Photosystem II (PSII) assembly is a stepwise process in which intermediate complexes with auxiliary proteins are transiently formed to allow efficient de novo biogenesis or repair of damaged PSII. In particular, the role of extrinsic PSII subunits (PsbO, PsbU, PsbV) and auxiliary proteins such as Psb27 for the formation and photoactivation of the Mn4O5Ca cluster, which catalyzes the unique water splitting reaction in mature PSII, remains unclear. Using cryo-electron microscopy, we have determined the structure of two novel late-stage PSII assembly intermediates from Thermosynechococcus vestitus BP-1. In contrast to previous studies, the resulting monomeric PSII complexes contain both PsbJ and Psb27 and exhibit a fully mature acceptor side, while the oxygen evolving complex (OEC) is still in an immature state. The second complex additionally associates with the late-acting assembly factor Psb32 and the extrinsic subunit PsbV. While Psb32 has received little attention, its proposed role in the complex challenges the previous assumption that all extrinsic subunits associate spontaneously, as well as the notion that PsbO initiates binding and solely drives OEC formation. Our structures of the Psb27-PSII and Psb32-PSII intermediates provide novel insights, how structural changes of C-termini of the D1 and D2 core proteins regulate maturation of the OEC and how the catalytic side is prepared for binding of the Mn4O5Ca cluster. The Psb32-PSII complex potentially represents the final PSII assembly intermediate that precedes the incorporation and photoactivation of the Mn4O5Ca cluster, allowing us to explain the final steps in the PSII biogenesis and assembly pipeline in great detail, as only the two extrinsic subunits PsbO and PsbU are missing.

plant biology↗

bioRxiv2024

Frequent transitions in self-assembly across the evolution of a central metabolic enzyme

Many enzymes assemble into homomeric protein complexes comprising multiple copies of one protein. Because structural form is usually assumed to follow function in biochemistry, these assemblies are thought to evolve because they provide some functional advantage. In many cases, however, no specific advantage is known and, in some cases, quaternary structure varies among orthologs. This has led to the proposition that self-assembly may instead vary neutrally within protein families. The extent of such variation has been difficult to ascertain because quaternary structure has until recently been difficult to measure on large scales. Here, we employ mass photometry, phylogenetics, and structural biology to interrogate the evolution of homo-oligomeric assembly across the entire phylogeny of prokaryotic citrate synthases - an enzyme with a highly conserved function. We discover a menagerie of different assembly types that come and go over the course of evolution, including cases of parallel evolution and reversions from complex to simple assemblies. Functional experiments in vitro and in vivo indicate that evolutionary transitions between different assemblies do not strongly influence enzyme catalysis. Our work suggests that enzymes can wander relatively freely through a large space of possible assemblies and demonstrates the power of characterizing structure-function relationships across entire phylogenies.

evolutionary biology↗

bioRxiv2024

Molecular principles of redox-coupled sodium pumping of the ancient Rnf machinery

The Rnf complex is the primary respiratory enzyme of several anaerobic prokaryotes that transfers electrons from ferredoxin to NAD+ and pumps sodium ions (Na+) across a membrane, powering ATP synthesis. Rnf is widespread in primordial organisms and the evolutionary predecessor of the Na+-pumping NADH-quinone oxidoreductase (Nqr)1. By running in reverse, Rnf reduces ferredoxin with NADH as reductant at the expense of the transmembrane electrochemical ion gradient and provides low potential electrons for nitrogenases as well as CO2 reductases. Yet, the molecular principles that couple the long-range electron transfer to the Na+ translocation across the membrane remain elusive. Here we resolve key functional states along the electron transfer pathway using redox-controlled cryo-electron microscopy (cryo-EM) that, in combination with biochemical functional assays and atomistic molecular simulations, provide key insight into the redox-driven Na+ pumping mechanism. We show that the reduction of the unique membrane-embedded [2Fe2S] cluster in the vestibule between the RnfA/E subunits electrostatically attracts the sodium ions, and in turn, triggers an inward/outward transition with alternating membrane access driving the Na+ pump and the reduction of NAD+. Our study unveils an ancient mechanism for redox-driven ion pumping, and provides key understanding of the fundamental principles governing energy conversion in biological systems.

biochemistry↗

bioRxiv2024

2-oxoglutarate triggers assembly of active dodecameric Methanosarcina mazei glutamine synthetase

Glutamine synthetases (GS) are central enzymes essential for the nitrogen metabolism across all domains of life. Consequently, they have been extensively studied for more than half a century. Based on the ATP dependent ammonium assimilation generating glutamine, GS expression and activity are strictly regulated in all organisms. In the methanogenic archaeon Methanosarcina mazei, it has been shown that the metabolite 2-oxoglutarate (2-OG) directly induces the GS activity. Besides, modulation of the activity by interaction with small proteins (GlnK1 and sP26) has been reported. Here, we show that the strong activation of M. mazei GS (GlnA1) by 2-OG is based on the 2-OG dependent dodecamer assembly of GlnA1 by using mass photometry (MP) and single particle cryo-electron microscopy (cryo-EM) analysis of purified strep-tagged GlnA1. The dodecamer assembly from dimers occurred without any detectable intermediate oligomeric state and was not affected in the presence of GlnK1. The 2.39 [A] cryo-EM structure of the dodecameric complex in the presence of 12.5 mM 2-OG demonstrated that 2-OG is binding between two monomers. Thereby, 2-OG appears to induce the dodecameric assembly in a cooperative way. Furthermore, the active site is primed by an allosteric interaction cascade caused by 2-OG-binding towards an adaption of an open active state conformation. In the presence of additional glutamine, strong feedback inhibition of GS activity was observed. Since glutamine dependent disassembly of the dodecamer was excluded by MP, feedback inhibition most likely relies on an allosteric binding of glutamine to the catalytic site. Based on our findings, we propose that under nitrogen limitation the induction of M. mazei GS into a catalytically active dodecamer is not affected by GlnK1 and crucially depends on the presence of 2-OG.

microbiology↗