We report the isolation and characterization of a novel nicotinic acetylcholine receptor (nAChR) ligand. The toxin is an 18 amino acid peptide and is the first reported α-conotoxin from an Atlantic fish-hunting Conus. The peptide was purified from the venom of Conus ermineus and is called a-conotoxin Ei. The sequence diverges from that of previously isolated α-conotoxins. We demonstrate that this structural divergence has functional consequences. In Torpedo nAChRs, α-conotoxin Ei selectively binds the agonist site near the aId subunit interface in contrast to α-conotoxin Mi which selectively targets the α/γ agonist binding site. In mammalian nAChRs α-conotoxin Ei shows high affinity for both the α/δ and α/γ subunit interfaces (with some preference for the α/δ site), whereas α-conotoxin Mi is highly selective for the α/δ ligand binding site. The sequence of the peptide is: Arg-Asp-Hyp-Cys-Cys-Tyr- His-Pro-Thr-Cys-Asn-Met-Ser-Asn-Pro-Gln-Ile-Cys-NH2, with disulfide bridging between Cys4-Cys10 and Cys5-Cys18, analogous to those of previously described α-conotoxins. This sequence has been verified by total chemical synthesis. Thus, α-conotoxin Ei is a newly-available tool with unique structure and function for characterization of nAChRs.
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