Differential targeting of nicotinic acetylcholine receptors by novel αA-conotoxins

We describe the isolation and characterization of two peptide toxins from Conus ermineus venom targeted to nicotinic acetylcholine receptors (nAChRs). The peptide structures have been confirmed by mass spectrometry and chemical synthesis. In contrast to the 12-18 residue, 4 Cys-containing α- conotoxins, the new toxins have 30 residues and 6 Cys residues. The toxins, named αA-conotoxins EIVA and EIVB, block both Torpedo and mouse α1- containing muscle subtype nAChRs expressed in Xenopus oocytes at low nanomolar concentrations. In contrast to α-bungarotoxin, αA-EIVA is inactive at α7-containing nAChRs even at micromolar concentrations. In this regard, αA-EIVA is similar to the previously described α-conotoxins (e.g. α-MI and α-GI) which also selectively target α1- versus α7-containing nAChRs. However, α-MI and α-GI discriminate between the α/δ versus α/γ subunit interfaces of the mouse muscle nAChR with 10,000-fold selectivity. In contrast, αA-conotoxin EIVA blocks both the α/γ site and α/δ site with equally high affinity but with distinct kinetics. The αA-conotoxins thus represent novel probes for the α/γ as well as the α/δ binding sites of the nAChR.