Biology subjects

Melling, O.

Publications and source records attributed to Melling, O..

4 recordsLinked to original sources

bioRxiv2026

Cryo-EM Structure of a Triazole alpha-Conotoxin GI Mimetic Bound to the Muscle-Type Nicotinic Acetylcholine Receptor

Disulfide-rich peptides possess exceptional potency and selectivity but are often limited by the instability and synthetic challenges associated with native disulfide bonds. Here, we report the design, synthesis, pharmacological evaluation, and structural characterisation of triazole-based peptidomimetics of the -GI conotoxin, a selective antagonist of the muscle-type nicotinic acetylcholine receptor (nAChR). A series of 1,4- and 1,5-disubstituted triazole analogues were prepared entirely on resin using CuAAC and RuAAC chemistry to replace the native Cys3/13 disulfide bridge. Functional evaluation against human muscle nAChRs revealed that 1,5-triazole analogues retained low-nanomolar potency, with the lead mimetic exhibiting activity comparable to native -GI. Cryo-electron microscopy of the lead compound bound to the muscle-type nAChR provided the first structure of a disulfide-isostere peptidomimetic in complex with a membrane receptor. The structure demonstrates that the 1,5-triazole reproduces the native peptide fold with high fidelity while contributing receptor-facing interactions not available to the native disulfide bridge. Molecular dynamics simulations further revealed conserved hydration networks and similar conformational sampling between the native peptide and lead mimetic. Together, these findings establish triazoles as effective disulfide surrogates and provide a structural framework for the rational design of stabilised conotoxin therapeutics.

biochemistry↗

bioRxiv2026

Structures of muscle-type nicotinic acetylcholine receptor with α-conotoxins reveal determinants of receptor-subtype specificity

-Conotoxins (-CTX) are disulfide-rich peptide antagonists with exceptional potency and subtype selectivity across nicotinic acetylcholine receptors (nAChRs), making them promising leads for therapeutic development. The molecular basis of their specificity has remained unresolved for decades, limiting rational drug design. Here, we present the first high-resolution cryo-EM structures of full-length muscle-type nAChR bound to three distinct -CTXs, revealing their precise binding modes at both acetylcholine binding sites. Our integrated approach, combining structural biology, pharmacological profiling, and computational hydration mapping, uncovers conserved pharmacophore features, water-mediated interactions, and subunit-specific determinants that govern potency and selectivity. These findings provide a long-sought molecular framework for -conotoxin recognition and inhibition, addressing a critical gap required to realise the full potential of conotoxins as lead compounds in drug discovery.

molecular biology↗

bioRxiv2025

Ligand-responsive groove remodelling in human and macaque CD1d reveals a conserved MHC-like gating mechanism

CD1d presents lipid antigens to invariant natural killer T (iNKT) cells. We determined a high-resolution crystal structure of human CD1d bound to -galactosylceramide (-GalCer) at 1.76 [A], enabling detailed investigation of ligand-sensitive conformational flexibility at Phe84, a conserved aromatic residue that caps the F' groove. Electron density at Phe84 revealed multiple side-chain conformations, suggestive of ligand-induced plasticity. Molecular dynamics simulations indicated that the canonical rotamer is energetically favoured in the absence of a stabilising groove-occupying ligand. To assess conservation of this putative gating mechanism, we solved the first CD1d structure from a non-human primate, rhesus macaque CD1d--GalCer, at 1.83 [A] resolution. In contrast to the human complex, Phe84 in macaque CD1d adopted a fixed conformation. As this aromatic residue is conserved across CD1 isoforms and CD1d-expressing species, and mirrors gating residues in MHC class I that regulate peptide accommodation, our findings support a shared evolutionary strategy for managing antigen diversity. These data provide critical insight into the mechanisms of antigen presentation by CD1 molecules. Significance StatementThis study reveals that Phe84, a conserved aromatic residue in CD1d, may act as a ligand-responsive gate modulating F' groove accessibility. This conditional plasticity could enable binding of structurally diverse lipid antigens and appears conserved across CD1 isoforms. The mechanism parallels class I MHC, where gating residues regulate peptide presentation, suggesting an evolutionarily shared strategy for accommodating antigen diversity.

immunology↗

bioRxiv2021

Interaction of the TonB dependent transporter HasR with its cognate TonB-like protein HasB in a membrane environment

The envelope of Gram-negative bacteria is composed of a double membrane separated by the periplasmic space. This organization imposes geometrical and distance constraints that are key for the mechanism of action of multicomponent systems spanning the envelope. However, consideration of all three compartments by experimental approaches is still elusive. Here we used the state-of-the-art molecular dynamics simulation in an Escherichia coli envelope model to obtain a dynamic view of molecular interactions between the outer membrane heme transporter HasR and the inner membrane TonB-like protein HasB. Their interaction allows the transfer of the inner membrane proton motive force derived energy to the transporter for heme internalization. The simulations which incorporate both membranes show the key role of periplasmic domains of both proteins, and their dynamics in the complex formation and stability. They revealed a previously unidentified atomic network of interactions, as well as the sequences of the interactions and their variations with the presence of external substrates. Experimental validation (mutations, phenotypic and in vitro assays) confirms the robustness of our approach and provides verification of the simulation-predicted interactions. Based on structural and sequence conservation, the network of interaction revealed in this study is expected to occur in other nutrient import systems. The integrative approach presented here is highly useful to study the dynamic interplay between components of any other bacterial transmembrane systems.

biophysics↗