NS6740 is an α7 nicotinic acetylcholine receptor-selective partial agonist with low efficacy for channel activation, capable of promoting the stable conversion of the receptors to nonconducting (desensitized) states that can be reactivated with the application of positive allosteric modulators (PAMs). In spite of its low efficacy for channel activation, NS6740 is an effective activator of the cholinergic anti-inflammatory pathway. We observed that the concentration-response relationships for channel activation, both when applied alone and when co-applied with the PAM PNU-120596 are inverted-U shaped with inhibitory/desensitizing activities dominant at high concentrations. We evaluated the potential importance of recently identified binding sites for allosteric activators and tested the hypotheses that the stable desensitization produced by NS6740 may be due to binding to these sites. Our experiments were guided by molecular modeling of NS6740 binding to both the allosteric and orthosteric activation sites on the receptor. Our results indicate that with α7C190A mutants, which have compromised orthosteric activation sites, NS6740 may work at the allosteric activation sites to promote transient PAM-dependent currents but not the stable desensitization seen with wild-type α7 receptors. Modeling NS6740 in the orthosteric binding sites identified S36 as an important residue for NS6740 binding and predicted that an S36V mutation would limit NS6740 activity. The efficacy of NS6740 for α7S36V receptors was reduced to zero, and applications of the compound to α7S36V receptors failed to induce the desensitization observed with wild-type receptors. The results indicate that the unique properties of NS6740 are due primarily to binding at the sites for orthosteric agonists.
Keywords: Allosteric activation; Docking analysis; Partial agonist; Silent agonist; Site-directed mutant; α(7) nicotinic acetylcholine receptor.
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