Interactions of Polyammonium Compounds with Isolated Acetylcholine Receptors

SCHMIDT and M. A. RAFTERY California Institute of Technology, Pasadena, Calif, U.S.A. 

Alan Rembaum and Eric Selegny (eds ), Polyelectrolytes and Their Applications, 175-185 All Rights Reserved Copyright 1975 by D Reidel Publishing Company, Dordrecht-Holland 

The techniques used include equilibrium dialysis [11] and fluorescence procedures [12]. Another method is based on the observation that the rate of binding of a-bungarotoxin (aBgt) to the receptor is affected by cholinergic ligands [13] and permits rapid estimation of the receptor affinities of a wide spectrum of nonradioactive, nondialyzable drugs. 

Torpedo AcChR was purified by an affinity chromatography procedure [II] DEAE-cellulose chromatography of the purified material, followed by concentration on a PM 10 dia-flow membrane yielded a product with a specific activity of 10 nmoles aBgt binding sites per mg protein (or 1 binding site per 100000 daltons of protein). 

Binding of iodotoxin to AcChR was monitored using a DEAE-cellulose disk technique described elsewhere [13] with the following modifications receptor concentration was reduced to 3 x 10 M concentration of toxin to 6x 10 M all experiments were carried out at 0° in 0.25 ml 2 mM sodium phosphate, pH 7.4,0.1% Triton X-100. Receptor was incubated with ligand for an hour formation of receptor-toxin complex was started by the addition of toxin and (unless otherwise stated) terminated after 0.20 min by applying an 0.1 ml aliquot to the DEAE-cellulose disk and immersing it in the wash fluid. Controls are obtained with receptor denatured by heating to 100° for 3 min. 

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INTERACTIONS OF POLYAMMONIUM COMPOUNDS WITH ISOLATED ACETYLCHOLINE RECEPTORS... 

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