Abstract
α-Conotoxins potently inhibit isoforms of nicotinic acetylcholine receptors (nAChRs), which are essential for neuronal and neuromuscular transmission. They are also used as neurochemical tools to study nAChR physiology and are being evaluated as drug leads to treat various neuronal disorders. A number of experimental studies have been performed to investigate the structure-activity relationships of conotoxin/nAChR complexes. However, the structural determinants of their binding interactions are still ambiguous in the absence of experimental structures of conotoxin-receptor complexes. In this study, the binding modes of α-conotoxin ImI to the α7-nAChR, currently the best-studied system experimentally, were investigated using comparative modeling and molecular dynamics simulations. The structures of more than 30 single point mutants of either the conotoxin or the receptor were modeled and analyzed. The models were used to explain qualitatively the change of affinities measured experimentally, including some nAChR positions located outside the binding site. Mutational energies were calculated using different methods that combine a conformational refinement procedure (minimization with a distance dependent dielectric constant or explicit water, or molecular dynamics using five restraint strategies) and a binding energy function (MM-GB/SA or MM-PB/SA). The protocol using explicit water energy minimization and MM-GB/SA gave the best correlations with experimental binding affinities, with an R2 value of 0.74. The van der Waals and non-polar desolvation components were found to be the main driving force for binding of the conotoxin to the nAChR. The electrostatic component was responsible for the selectivity of the various ImI mutants. Overall, this study provides novel insights into the binding mechanism of α-conotoxins to nAChRs and the methodological developments reported here open avenues for computational scanning studies of a rapidly expanding range of wild-type and chemically modified α-conotoxins.
Highlights
Nicotinic acetylcholine receptors are a large family of ligand-gated ion channels that mediate rapid synaptic transmission in the central and peripheral nervous system [1,2]. nicotinic acetylcholine receptors (nAChRs) are implicated in disorders such as Alzheimer’s diseases, schizophrenia, depression, hyperactivity disorders and tobacco addiction [3,4,5,6]
All nAChRs are comprised of five homologous subunits, which are divided into a large N-terminal extracellular ligand-binding domain (LBD), a transmembrane domain, and an intracellular domain [7] (Figure 1)
The aim of our study is to compare different methods to predict the impact of single point mutations on binding affinities between conotoxin ImI and a7-nAChR; the use of unnatural residues would complicate the interpretation of those comparisons as the deviation between computed and experimental mutational energies could arise from inaccuracy in the parameters as well as from the methodological differences
Summary
Nicotinic acetylcholine receptors (nAChRs) are a large family of ligand-gated ion channels that mediate rapid synaptic transmission in the central and peripheral nervous system [1,2]. nAChRs are implicated in disorders such as Alzheimer’s diseases, schizophrenia, depression, hyperactivity disorders and tobacco addiction [3,4,5,6]. Nicotinic acetylcholine receptors (nAChRs) are a large family of ligand-gated ion channels that mediate rapid synaptic transmission in the central and peripheral nervous system [1,2]. The nAChR subtypes include hetero- or homo-pentamers of a1-10, c, b1-4, d and/or e subunits. These subtypes differ in their pharmacological and kinetic properties, as well as their localization [8,9]. This topology includes four cysteines arranged in a common sequence pattern -CCXmCXnC-, where X is any non-cysteine residue, and n and m are the numbers of intercysteine residues. Disulfide bonds connect cysteines I-III and II-IV [18,19]
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