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Diversity of Nicotinic Acetylcholine Receptor Positive Allosteric Modulators Revealed by Mutagenesis and a Revised Structural Model.

ABSTRACT: By combining electrophysiological and computational approaches we have examined a series of positive allosteric modulators (PAMs) acting on the human ?7 nicotinic acetylcholine receptor (nAChR). Electrophysiological studies have focused on three ?7-selective PAMs (A-867744, TBS-516, and TQS) that display similar effects on wild-type ?7 nAChRs. In addition to potentiating agonist-evoked responses, all three compounds reduce receptor desensitization and, consequently, are classed as type II PAMs. Despite having similar effects on wild-type receptors, A-867744 was found to have profoundly differing effects on mutated receptors compared with TBS-516 and TQS, a finding that is consistent with previous studies indicating that A-867744 may have a different mechanism of action compare with other ?7-selective type II PAMs. Due to evidence that these PAMs bind within the ?7 nAChR transmembrane region, we generated and validated new structural models of ?7. Importantly, we have corrected a previously identified error in the transmembrane region of the original cryo-electron microscopy Torpedo model; the only pentameric ligand-gated ion channel imaged in a native lipid membrane. Real-space refinement was used to generate closed and open conformations on which the ?7 models were based. Consensus docking with an extended series of PAMs with chemical similarity to A-867744, TBS-516, and TQS suggests that all bind to a broadly similar intersubunit transmembrane site. However, differences in the predicted binding of A-867744, compared with TBS-516 and TQS, may help to explain the distinct functional effects of A-867744. Thus, our revised structural models may provide a useful tool for interpreting functional effects of PAMs.

SUBMITTER: Newcombe J

PROVIDER: S-EPMC5767682 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

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Diversity of Nicotinic Acetylcholine Receptor Positive Allosteric Modulators Revealed by Mutagenesis and a Revised Structural Model.

Newcombe Joseph J Chatzidaki Anna A Sheppard Tom D TD Topf Maya M Millar Neil S NS

Molecular pharmacology 20171201 2

By combining electrophysiological and computational approaches we have examined a series of positive allosteric modulators (PAMs) acting on the human α7 nicotinic acetylcholine receptor (nAChR). Electrophysiological studies have focused on three α7-selective PAMs (A-867744, TBS-516, and TQS) that display similar effects on wild-type α7 nAChRs. In addition to potentiating agonist-evoked responses, all three compounds reduce receptor desensitization and, consequently, are clas ...[more]

PMID: 29196491

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Project description:Homomeric α7 nicotinic acetylcholine receptors (nAChR) have an intrinsically low probability of opening that can be overcome by α7-selective positive allosteric modulators (PAMs), which bind at a site involving the second transmembrane domain (TM2). Mutation of a methionine that is unique to α7 at the 15' position of TM2 to leucine, the residue in most other nAChR subunits, largely eliminates the activity of such PAMs. We tested the effect of the reverse mutation (L15'M) in heteromeric nAChR receptors containing α4 and β2, which are the nAChR subunits that are most abundant in the brain. Receptors containing these mutations were found to be strongly potentiated by the α7 PAM 3a,4,5,9b-tetrahydro-4-(1-naphthalenyl)-3H-cyclopentan[c]quinoline-8-sulfonamide (TQS) but insensitive to the alternative PAM 1-(5-chloro-2,4-dimethoxyphenyl)-3-(5-methylisoxazol-3-yl)-urea. The presence of the mutation in the β2 subunit was necessary and sufficient for TQS sensitivity. The primary effect of the mutation in the α4 subunit was to reduce responses to acetylcholine applied alone. Sensitivity to TQS required only a single mutant β subunit, regardless of the position of the mutant β subunit within the pentameric complex. Similar results were obtained when β2L15'M was coexpressed with α2 or α3 and when the L15'M mutation was placed in β4 and coexpressed with α2, α3, or α4. Functional receptors were not observed when β1L15'M subunits were coexpressed with other muscle nAChR subunits. The unique structure-activity relationship of PAMs and the α4β2L15'M receptor compared with α7 and the availability of high-resolution α4β2 structures may provide new insights into the fundamental mechanisms of nAChR allosteric potentiation. SIGNIFICANCE STATEMENT: Heteromeric neuronal nAChRs have a relatively high initial probability of channel activation compared to receptors that are homomers of α7 subunits but are insensitive to PAMs, which greatly increase the open probability of α7 receptors. These features of heteromeric nAChR can be reversed by mutation of a single residue present in all neuronal heteromeric nAChR subunits to the sequence found in α7. Specifically, the mutation of the TM2 15' leucine to methionine in α subunits reduces heteromeric receptor channel activation, while the same mutation in neuronal β subunits allows heteromeric receptors to respond to select α7 PAMs. The results indicate a key role for this residue in the functional differences in the two main classes of neuronal nAChRs.

Dataset Information

Diversity of Nicotinic Acetylcholine Receptor Positive Allosteric Modulators Revealed by Mutagenesis and a Revised Structural Model.

Publications

Diversity of Nicotinic Acetylcholine Receptor Positive Allosteric Modulators Revealed by Mutagenesis and a Revised Structural Model.

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