Differential targeting of nicotinic acetylcholine receptors by novel alpha A-conotoxins

We describe the isolation and characterization of two peptide toxins from Conus ermimeus 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 alpha-conotoxins, the new toxins have 30 residues and 6 Cys residues. The toxins, named alpha A-conotoxins EIVA and EIVB, block both Torpedo and mouse alpha 1-containing muscle subtype nAChRs expressed in Xenopus oocytes at low nanomolar concentrations. In contrast to alpha-bungarotoxin, alpha A-EIVA is inactive at alpha 7-containing nAChRs even at micromolar concentrations. In this regard, alpha A-EIVA is similar to the previously described alpha-conotoxins (e.g. alpha-MI and alpha-GI) which also selectively target alpha 1- versus alpha 7-containing nAChRs, However, alpha-MI and alpha-GI discriminate between the alpha/delta versus alpha/gamma subunit interfaces of the mouse muscle nAChR with 10,000-fold selectivity. In contrast, alpha A-conotoxin EIVA blocks both the alpha/gamma site and alpha/delta site with equally high affinity but with distinct kinetics. The alpha A-conotoxins thus represent novel probes for the alpha/gamma as well as the alpha/delta binding sites of the nAChR.