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Acetylcholine Promotes Binding of α-Conotoxin MII at α3β2 Nicotinic Acetylcholine Receptors

Authors

  • Somisetti V. Sambasivarao,

    1. Chemical and Biological Engineering Department, Colorado School of Mines, 1500 Illinois Street, Golden, CO 80401 (USA)
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  • Jessica Roberts,

    1. Department of Biological Sciences, Idaho State University, 650 Memorial Drive, Pocatello, ID 83209 (USA)
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  • Vivek S. Bharadwaj,

    1. Chemical and Biological Engineering Department, Colorado School of Mines, 1500 Illinois Street, Golden, CO 80401 (USA)
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  • Jason G. Slingsby,

    1. Chemical and Biological Engineering Department, Colorado School of Mines, 1500 Illinois Street, Golden, CO 80401 (USA)
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  • Conrad Rohleder,

    1. Chemical and Biological Engineering Department, Colorado School of Mines, 1500 Illinois Street, Golden, CO 80401 (USA)
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  • Chris Mallory,

    1. Department of Chemistry and Biochemistry, Boise State University, 1910 University Drive, Boise, ID 83725 (USA)
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  • Prof. James R. Groome,

    1. Department of Biological Sciences, Idaho State University, 650 Memorial Drive, Pocatello, ID 83209 (USA)
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  • Prof. Owen M. McDougal,

    Corresponding author
    1. Department of Chemistry and Biochemistry, Boise State University, 1910 University Drive, Boise, ID 83725 (USA)
    • Owen M. McDougal, Department of Chemistry and Biochemistry, Boise State University, 1910 University Drive, Boise, ID 83725 (USA)

      C. Mark Maupin, Chemical and Biological Engineering Department, Colorado School of Mines, 1500 Illinois Street, Golden, CO 80401 (USA)

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  • Prof. C. Mark Maupin

    Corresponding author
    1. Chemical and Biological Engineering Department, Colorado School of Mines, 1500 Illinois Street, Golden, CO 80401 (USA)
    • Owen M. McDougal, Department of Chemistry and Biochemistry, Boise State University, 1910 University Drive, Boise, ID 83725 (USA)

      C. Mark Maupin, Chemical and Biological Engineering Department, Colorado School of Mines, 1500 Illinois Street, Golden, CO 80401 (USA)

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Abstract

α-Conotoxin MII (α-CTxMII) is a 16-residue peptide with the sequence GCCSNPVCHLEHSNLC, containing Cys2–Cys8 and Cys3–Cys16 disulfide bonds. This peptide, isolated from the venom of the marine cone snail Conus magus, is a potent and selective antagonist of neuronal nicotinic acetylcholine receptors (nAChRs). To evaluate the impact of channel–ligand interactions on ligand-binding affinity, homology models of the heteropentameric α3β2-nAChR were constructed. The models were created in MODELLER with the aid of experimentally characterized structures of the Torpedo marmorata-nAChR (Tm-nAChR, PDB ID: 2BG9) and the Aplysia californica-acetylcholine binding protein (Ac-AChBP, PDB ID: 2BR8) as templates for the α3- and β2-subunit isoforms derived from rat neuronal nAChR primary amino acid sequences. Molecular docking calculations were performed with AutoDock to evaluate interactions of the heteropentameric nAChR homology models with the ligands acetylcholine (ACh) and α-CTxMII. The nAChR homology models described here bind ACh with binding energies commensurate with those of previously reported systems, and identify critical interactions that facilitate both ACh and α-CTxMII ligand binding. The docking calculations revealed an increased binding affinity of the α3β2-nAChR for α-CTxMII with ACh bound to the receptor, and this was confirmed through two-electrode voltage clamp experiments on oocytes from Xenopus laevis. These findings provide insights into the inhibition and mechanism of electrostatically driven antagonist properties of the α-CTxMIIs on nAChRs.

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