title: Ligand binding to nicotinic acetylcholine receptor investigated by surface plasmon resonance.
abstract: Ligand binding to the nicotinic acetylcholine receptor is studied by surface plasmon resonance. Biotinylated bungarotoxin, immobilized on a streptavidin-coated gold film, binds nicotinic acetylcholine receptor both in detergent-solubilized and in lipid vesicle-reconstituted form with high specificity. In the latter case, nonspecific binding to the sensor surface is significantly reduced by reconstituting the receptor into poly(ethylene glycol)-lipid-containing sterically stabilized vesicles. By preincubation of a bulk nicotinic acetylcholine receptor sample with the competing ligands carbamoylcholine and decamethonium bromide, the subsequent specific binding of the receptor to the surface-immobilized bungarotoxin is reduced, depending on the concentration of competing ligand. This competition assay allows the determination of the dissociation constants of the acetylcholine receptor-carbamoylcholine complex. A K(D) = 3.5 × 10(-)(6) M for the detergent-solubilized receptor and a K(D) = 1.4 × 10(-)(5) M for the lipid vesicle-reconstituted receptor are obtained. For decamethonium bromide, a K(D) = 4.5 × 10(-)(5) M is determined for the detergent-solubilized receptor. This approach is of general importance for investigating ligand-receptor interactions in case of small ligand molecules by mass-sensitive techniques.
authors.affiliation: Laboratoire de Chimie Physique des Polymères et Membranes, Ecole Polytechnique Fédérale de Lausanne, CH-1015 Lausanne, Switzerland, and Institut für Biochemie, Freie Universität Berlin, Thielallee 63, D-14195 Berlin.
authors.forename: D
authors.initials: D
authors.lastname: Kröger
authors.forename: F
authors.initials: F
authors.lastname: Hucho
authors.forename: H
authors.initials: H
authors.lastname: Vogel