Activation and Desensitization of Torpedo Acetylcholine Receptor: Evidence for Separate Binding Sites

Overview

Journal Proc Natl Acad Sci U S A

Specialty Science

Date 1982 Nov 1

PMID 6960348

Citations 12

Authors

S M Dunn

M A RAFTERY

Affiliations

Soon will be listed here.

Abstract

The acetylcholine receptor from Torpedo californica was labeled by reaction with the fluorescent probe 4-[N-(iodoacetoxy)ethyl-N-methyl]amino-7-nitrobenz-2-oxa-1,3-diazole without apparent effect on its in vitro ligand binding and functional properties. Addition of acetylcholine or carbamoylcholine to the labeled-receptor preparations enhanced the fluorescence of the bound probe, and this effect was specific for agonists and inhibited by prior incubation with excess alpha-bungarotoxin. Equilibrium fluorescence titrations gave apparent dissociation constants of 0.86 +/- 0.14 mM for carbamoylcholine and 79 +/- 11 microM for acetylcholine, in good agreement with the dissociation constants measured for the permeability response of the receptor. Stopped-flow experiments showed that the fluorescence change was a single exponential process whose rate increased with ligand concentration, reaching a saturating value for carbamoylcholine of approximately 400 s-1. The equilibrium binding of carbamoylcholine was not significantly affected by prior incubation of the receptor with d-tubocurarine or histrionicotoxin and the dissociation constant was only slightly increased in the presence of lidocaine. These inhibitory ligands do not, therefore, compete directly with agonists for this low-affinity binding site, suggesting that their mode of action may be indirect.

Citing Articles

Kinetics of agonist-induced intrinsic fluorescence changes in the Torpedo acetylcholine receptor.

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Epibatidine binds to four sites on the Torpedo nicotinic acetylcholine receptor.

Kawai H, Dunn S, Raftery M Biochem Biophys Res Commun. 2007; 366(3):834-9.

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Photoaffinity labeling of Torpedo acetylcholine receptor at multiple sites.

Tine S, RAFTERY M Proc Natl Acad Sci U S A. 1993; 90(15):7308-11.

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Time-resolved photolabeling by the noncompetitive blocker chlorpromazine of the acetylcholine receptor in its transiently open and closed ion channel conformations.

Heidmann T, Changeux J Proc Natl Acad Sci U S A. 1984; 81(6):1897-901.

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Brain and muscle nicotinic acetylcholine receptors are different but homologous proteins.

Dunn S, Barnard E, Dolly J, Lai F, Ray N, RAFTERY M Proc Natl Acad Sci U S A. 1985; 82(15):5208-12.

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