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Abstract The Mitochondrial Complex I Assembly (MCIA) complex is essential for the biogenesis of respiratory Complex I (CI), the first enzyme in the respiratory chain, which has been linked to Alzheimer’s disease (AD) pathogenesis. However, how MCIA facilitates CI assembly, and how it is linked with AD pathogenesis, is poorly understood. Here we report the structural basis of the complex formation between the MCIA subunits ECSIT and ACAD9. ECSIT binding induces a major conformational change in the FAD-binding loop of ACAD9, releasing the FAD cofactor and converting ACAD9 from a fatty acid β-oxidation (FAO) enzyme to a CI assembly factor. We provide evidence that ECSIT phosphorylation downregulates its association with ACAD9 and is reduced in neuronal cells upon exposure to amyloid-β (Aβ) oligomers. These findings advance our understanding of the MCIA complex assembly and suggest a possible role for ECSIT in the reprogramming of bioenergetic pathways linked to Aβ toxicity, a hallmark of AD.

Chlamydomonas reinhardtii Photosystem II (PSII) is a dimer consisting of at least 13 nuclear-encoded and four chloroplast-encoded protein subunits that collectively function as a sunlight-driven oxidoreductase. In this study, we present the inaugural structure of a green alga PSII assembly intermediate (pre-PSII-int). This intermediate was isolated from chloroplast membranes of the temperature-sensitive mutant TSP4, cultivated for 14 hours at a non-permissive temperature. The assembly state comprises a monomer containing subunits A, B, C, D, E, F, H, I, K, and two novel assembly factors, Psb1 and Psb2. Psb1 is identified as a novel transmembrane helix located adjacent to PsbE and PsbF (cytochrome b559). The absence of PsbJ, typically found in mature PSII close to this position, indicates that Psb1 functions as an assembly factor. Psb2 is an eukaryotic homolog of the cyanobacterial assembly factor Psb27. The presence of iron, coupled with the absence of QA, QB, and the manganese cluster, implies a protective mechanism against photodamage and provides insights into the intricate assembly process.

Abstract Multi-specific antibodies (msAbs) are being developed as next generation antibody-based therapeutics. Knowledge of the three-dimensional structures, in the full antibody context, of their fragment antigen-binding (Fab) moieties with or without bound antigens is key to elucidating their therapeutic efficiency and stability. However, the flexibility of msAbs, a feature essential for their multi specificity, has hindered efforts in this direction. Cross-Over Dual Variable immunoglobulin (CODVIg) is a promising bispecific antibody format, designed to simultaneously target the interleukins IL4 and IL13. In this work we present the biophysical and structural characterisation of a CODVFab:IL13 complex in the full antibody context, using cryo-electron microscopy at an overall resolution of 4.2 Å. Unlike the 1:2 stoichiometry previously observed for CODVIg:IL4, CODVIg:IL13 shows a 1:1 stoichiometry. As well as providing details of the IL13-CODV binding interface, including the residues involved in the epitope-paratope region, the structure of CODVFab:IL13 also validates the use of labelling antibody as a new strategy for the single particle cryo-EM study of msAbs in complex with one, or more, antigens. This strategy reduced the inherent flexibility of the IL13 binding domain of CODV without inducing either structural changes at the epitope level or steric hindrance between the IL4 and IL13 binding regions of CODVIg. The work presented here thus also contributes to the development of methodology for the structural study of msAbs, a promising platform for cancer immunotherapy.

Abstract To save energy and precisely regulate cardiac contractility, cardiac muscle myosin heads are sequestered in an ‘off’ state that can be converted to an ‘on’ state when exertion is increased. The ‘off’ state is equated with a folded-back structure known as the interacting-heads motif (IHM), which is a regulatory feature of all class-2 muscle and non-muscle myosins. We report here the human β-cardiac myosin IHM structure determined by cryo-electron microscopy to 3.6 Å resolution, providing details of all the interfaces stabilizing the ‘off’ state. The structure shows that these interfaces are hot spots of hypertrophic cardiomyopathy mutations that are thought to cause hypercontractility by destabilizing the ‘off’ state. Importantly, the cardiac and smooth muscle myosin IHM structures dramatically differ, providing structural evidence for the divergent physiological regulation of these muscle types. The cardiac IHM structure will facilitate development of clinically useful new molecules that modulate IHM stability.

Membrane lipid composition is a key factor for bacterial stress adaptation. Here, authors show that ViaA and AAA+ ATPase RavA, alter lipid composition and modulate E. coli aminoglycoside sensitivity.

Here the authors report the cryo-EM structure of a triple-mutant of the anti-microbial peptide plectasin, PPI42, assembling in a pH- and concentration dependent manner into helical non-amyloid fibrils. The fibrils formation is reversible, and follows a sigmoidal kinetics. The fibrils adopt a right-handed helical superstructure composed by two protofilaments, stabilized by an outer hydrophobic ring and an inner hydrophobic centre. These findings reveal that α/β proteins can natively assemble into fibrils.