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

SUMMARY α-Conotoxins (α-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 α-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 α-conotoxin recognition and inhibition, addressing a critical gap required to realise the full potential of conotoxins as lead compounds in drug discovery.