The Allosteric Activation Of Alpha 7 Nachr By Alpha-Conotoxin Mric Is Modified By Mutations At The Vestibular Site

alpha-conotoxins are 13-19 amino acid toxin peptides that bind various nicotinic acetylcholine receptor (nAChR) subtypes. alpha-conotoxin Mr1.7c (MrIC) is a 17 amino acid peptide that targets alpha 7 nAChR. Although MrIC has no activating effect on alpha 7 nAChR when applied by itself, it evokes a large response when co-applied with the type II positive allosteric modulator PNU-120596, which potentiates the alpha 7 nAChR response by recovering it from a desensitized state. A lack of standalone activity, despite activation upon co-application with a positive allosteric modulator, was previously observed for molecules that bind to an extracellular domain allosteric activation (AA) site at the vestibule of the receptor. We hypothesized that MrIC may activate alpha 7 nAChR allosterically through this site. We ran voltage-clamp electrophysiology experiments and in silico peptide docking calculations in order to gather evidence in support of alpha 7 nAChR activation by MrIC through the AA site. The experiments with the wild-type alpha 7 nAChR supported an allosteric mode of action, which was confirmed by the significantly increased MrIC + PNU-120596 responses of three alpha 7 nAChR AA site mutants that were designed in silico to improve MrIC binding. Overall, our results shed light on the allosteric activation of alpha 7 nAChR by MrIC and suggest the involvement of the AA site.