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Heterologous expression and nonsense suppression provide insights into agonist behavior at α6β2 nicotinic acetylcholine receptors

Post, Michael R. and Limapichat, Walrati and Lester, Henry A. and Dougherty, Dennis A. (2015) Heterologous expression and nonsense suppression provide insights into agonist behavior at α6β2 nicotinic acetylcholine receptors. Neuropharmacology, 97 . pp. 376-382. ISSN 0028-3908. PMCID PMC4635625. doi:10.1016/j.neuropharm.2015.04.009. https://resolver.caltech.edu/CaltechAUTHORS:20150429-110737552

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Abstract

The α6-containing subtypes of the nicotinic acetylcholine receptor (nAChR) are localized to presynaptic terminals of the dopaminergic pathways of the central nervous system. Selective ligands for these nAChRs are potentially useful in both Parkinson's disease and addiction. For these and other goals, it is important to distinguish the binding behavior of agonists at the α6-β2 binding site versus other subtypes. To study this problem, we apply nonsense suppression-based non-canonical amino acid mutagenesis. We report a combination of four mutations in α6β2 that yield high-level heterologous expression in Xenopus oocytes. By varying mRNA injection ratios, two populations were observed with unique characteristics, likely due to differing stoichiometries. Responses to nine known nAChR agonists were analyzed at the receptor, and their corresponding EC50 values and efficacies are reported. The system is compatible with nonsense suppression, allowing structure–function studies between Trp149 – a conserved residue on loop B found to make a cation-π interaction at several nAChR subtypes – and several agonists. These studies reveal that acetylcholine forms a strong cation-π interaction with the conserved tryptophan, while nicotine and TC299423 do not, suggesting a unique pharmacology for the α6β2 nAChR.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1016/j.neuropharm.2015.04.009DOIArticle
http://www.sciencedirect.com/science/article/pii/S0028390815001380PublisherArticle
https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4635625PubMed CentralArticle
ORCID:
AuthorORCID
Post, Michael R.0000-0002-3214-7619
Lester, Henry A.0000-0002-5470-5255
Dougherty, Dennis A.0000-0003-1464-2461
Additional Information:© 2015 Published by Elsevier Ltd. Received 14 January 2015; Received in revised form 27 March 2015; Accepted 10 April 2015; Available online 20 April 2015. We thank the NIH (NS 34407) for support of this work. MRP was supported by an NIH/NRSA training grant: 5 T32 GM07616.
Funders:
Funding AgencyGrant Number
NIHNS 34407
NIH/NRSA Training Grant5 T32 GM07616
Subject Keywords:Parkinson's disease; Addiction; Ion channels; Nicotinic acetylcholine receptors; Electrophysiology; Non-canonical amino acids
PubMed Central ID:PMC4635625
DOI:10.1016/j.neuropharm.2015.04.009
Record Number:CaltechAUTHORS:20150429-110737552
Persistent URL:https://resolver.caltech.edu/CaltechAUTHORS:20150429-110737552
Official Citation:Michael R. Post, Walrati Limapichat, Henry A. Lester, Dennis A. Dougherty, Heterologous expression and nonsense suppression provide insights into agonist behavior at α6β2 nicotinic acetylcholine receptors, Neuropharmacology, Volume 97, October 2015, Pages 376-382, ISSN 0028-3908, http://dx.doi.org/10.1016/j.neuropharm.2015.04.009. (http://www.sciencedirect.com/science/article/pii/S0028390815001380)
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:57088
Collection:CaltechAUTHORS
Deposited By: Tony Diaz
Deposited On:29 Apr 2015 20:04
Last Modified:10 Nov 2021 21:07

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