A characterization of α-bungarotoxin binding in the brain of the horseshoe crab, Limulus polyphemus

The binding of 125I-labeled α-bungarotoxin to membrane fragments prepared from Limulus brain tissue has been investigated. Toxin binding approaches saturation in the range of 30 to 40 n m, with maximum binding of 2 to 6 pmol/mg of protein. The saturation kinetics and the rate of displacement of bound toxin are consistent with multiple toxin binding sites. Pharmacological studies show that binding is inhibited by both cholinergic agonists and antagonists, I 50′s for inhibition by d-tubocurarine, nicotine, decamethonium, carbachol, and atropine are 2 × 10 −6, 7 × 10 −6, 2 × 10 −5, 6 × 10 −4, and 3 × 10 −4 m, respectively. Nicotinic ligands inhibited binding much more effectively than muscarinic ligands. Toxin binding activity was solubilized with Triton X-100. Velocity sedimentation analysis of the solubilized activity revealed three separate components. Seventy to eighty percent of the binding activity had a sedimentation coefficient of 8.6 S. The remaining activity was composed of two components with sedimentation coefficients of 15.1 and 17 S.