Despite the odds: formation of the SARS-CoV-2 methylation complex.

Coronaviruses modify their single-stranded RNA genome with a methylated cap during replication to mimic the eukaryotic mRNAs. The capping process is initiated by several nonstructural proteins (nsp) encoded in the viral genome. The methylation is performed by two methyltransferases, nsp14 and nsp16, while nsp10 acts as a co-factor to both. Additionally, nsp14 carries an exonuclease domain which operates in the proofreading system during RNA replication of the viral genome. Both nsp14 and nsp16 were reported to independently bind nsp10, but the available structural information suggests that the concomitant interaction between these three proteins would be impossible due to steric clashes. Here, we show that nsp14, nsp10, and nsp16 can form a heterotrimer complex upon significant allosteric change. This interaction is expected to encourage the formation of mature capped viral mRNA, modulating nsp14's exonuclease activity, and protecting the viral RNA. Our findings show that nsp14 is amenable to allosteric regulation and may serve as a novel target for therapeutic approaches.

Crystal structure of glycerol kinase from Trypanosoma cruzi, a potential molecular target in Chagas disease.

Chagas disease is a neglected tropical disease caused by the protozoan parasite Trypanosoma cruzi. It bears a significant global health burden with limited treatment options, thus calling for the development of new and effective drugs. Certain trypanosomal metabolic enzymes have been suggested to be druggable and valid for subsequent inhibition. In this study, the crystal structure of glycerol kinase from T. cruzi, a key enzyme in glycerol metabolism in this parasite, is presented. Structural analysis allowed a detailed description of the glycerol binding pocket, while comparative assessment pinpointed a potential regulatory site which may serve as a target for selective inhibition. These findings advance the understanding of glycerol metabolism in eukaryotes and provide a solid basis for the future treatment of Chagas disease.

Cryo-EM structure of flagellotropic bacteriophage Chi.

The flagellotropic bacteriophage χ (Chi) infects bacteria via the flagellar filament. Despite years of study, its structural architecture remains partly characterized. Through cryo-EM, we unveil χ's nearly complete structure, encompassing capsid, neck, tail, and tail tip. While the capsid and tail resemble phage YSD1, the neck and tail tip reveal new proteins and their arrangement. The neck shows a unique conformation of the tail tube protein, forming a socket-like structure for attachment to the neck. The tail tip comprises four proteins, including distal tail protein (DTP), two baseplate hub proteins (BH1P and BH2P), and tail tip assembly protein (TAP) exhibiting minimal organization compared to other siphophages. Deviating from the consensus in other siphophages, DTP in χ forms a trimeric assembly, reducing tail symmetry from 6-fold to 3-fold at the tip. These findings illuminate the previously unexplored structural organization of χ's neck and tail tip.

Atomic structures of naphthalene dipeptide micelles unravel mechanisms of assembly and gelation.

Peptide-based biopolymers have gained increasing attention due to their versatile applications. A naphthalene dipeptide (2NapFF) can form chirality-dependent tubular micelles, leading to supramolecular gels. The precise molecular arrangement within these micelles and the mechanism governing gelation have remained enigmatic. We determined, at near-atomic resolution, cryoelectron microscopy structures of the 2NapFF micelles LL-tube and LD-tube, generated by the stereoisomers (l,l)-2NapFF and (l,d)-2NapFF, respectively. The structures reveal that the fundamental packing of dipeptides is driven by the systematic π-π stacking of aromatic rings and that same-charge repulsion between the carbonyl groups is responsible for the stiffness of both tubes. The structural analysis elucidates how a single residue's altered chirality gives rise to markedly distinct tubular structures and sheds light on the mechanisms underlying the pH-dependent gelation of LL- and LD-tubes. The understanding of dipeptide packing and gelation mechanisms provides insights for the rational design of 2NapFF derivatives, enabling the modulation of micellar dimensions.

An extensive disulfide bond network prevents tail contraction in Agrobacterium tumefaciens phage Milano.

A contractile sheath and rigid tube assembly is a widespread apparatus used by bacteriophages, tailocins, and the bacterial type VI secretion system to penetrate cell membranes. In this mechanism, contraction of an external sheath powers the motion of an inner tube through the membrane. The structure, energetics, and mechanism of the machinery imply rigidity and straightness. The contractile tail of Agrobacterium tumefaciens bacteriophage Milano is flexible and bent to varying degrees, which sets it apart from other contractile tail-like systems. Here, we report structures of the Milano tail including the sheath-tube complex, baseplate, and putative receptor-binding proteins. The flexible-to-rigid transformation of the Milano tail upon contraction can be explained by unique electrostatic properties of the tail tube and sheath. All components of the Milano tail, including sheath subunits, are crosslinked by disulfides, some of which must be reduced for contraction to occur. The putative receptor-binding complex of Milano contains a tailspike, a tail fiber, and at least two small proteins that form a garland around the distal ends of the tailspikes and tail fibers. Despite being flagellotropic, Milano lacks thread-like tail filaments that can wrap around the flagellum, and is thus likely to employ a different binding mechanism.