Determination of the α-Conotoxin Vc1.1 Binding Site on the α9α10 Nicotinic Acetylcholine Receptor

Abstract

α-Conotoxin Vc1.1 specifically and potently inhibits the nicotinic acetylcholine receptor subtype α9α10 (α9α10 nAChR) and is a potential novel treatment for neuropathic pain. Here, we used a combination of computational modeling and electrophysiology experiments to determine the Vc1.1 binding site on the α9α10 nAChR. Interactions of Vc1.1 with two probable binding sites, α9α10 and α10α9, were modeled. Mutational energies calculated by assuming specific interactions in the α10α9 binding site correlated better with electrophysiological recordings than those assuming interactions with the α9α10 binding site. Two novel Vc1.1 analogues, [N9F]Vc1.1 and [N9W]Vc1.1, were predicted to have large differences in affinity between the two binding sites. Data from functional studies were consistent with computational predictions that assumed preferred binding of Vc1.1 to the α10α9 pocket. Moreover, our modeling study suggested that a single hydrogen bond formed between Vc1.1 and position 59 of the α10α9 pocket confers specificity to rat versus human α9α10 nAChRs.

Molecular Interaction of α-Conotoxin RgIA with the Rat α9α10 Nicotinic Acetylcholine Receptor

“…8). This a10(1)/a9(2) interface is the high-affinity binding site for a-CTx RgIA binding and consistent with a recent study, which also suggested a similar interaction for a-CTx Vc1.1 with the a10(1)/a9(2) interface of the rat a9a10 nAChR (Yu et al, 2013). Conotoxins that bind the a9a10 nAChR are being considered as potential pain therapeutics (McIntosh et al, 2009;Del Bufalo et al, 2014;Di Cesare Mannelli et al, 2014), and understanding the subunit determinants of a-CTx interaction with the a9a10 nAChR may facilitate further development of such compounds.…”

Identifying Key Amino Acid Residues That Affect α-Conotoxin AuIB Inhibition of α3β4 Nicotinic Acetylcholine Receptors

“…AuIB could also bind at the non-canonical binding site (ϩ)␤/ (Ϫ)␣, as suggested for galanthamine and cocaine binding to AChBP (65), anti-helminthic compound morantel binding to ␣3␤2 nAChRs (66), and recently for ␣-conotoxin Vc1.1 binding to ␣9␣10 nAChRs (67). However, this is unlikely because the docking model does not support binding at a non-canonical site and the [W59A]␤4 mutant loses inhibition despite identical WLK pockets being available on the ␣3 subunit (Ϫ) side.…”

Key Residues in the Nicotinic Acetylcholine Receptor β2 Subunit Contribute to α-Conotoxin LvIA Binding

“…The molecular model was refined by a 30-ns explicit water molecular dynamics simulation carried out with the GROMACS 4.6.5 (26) package and the ff03 force field (27), using a procedure described previously (28,29). All the models of complexes involving ␤2 subunit mutants were generated by substituting residue side chains using Modeler 9v14 (30).…”