Interactions between discrete neuronal membrane binding sites for the putative K + ‐channel ligands β‐bungarotoxin, dendrotoxin and mast‐cell‐degranulating peptide

1 β‐Bungarotoxin, a presynaptically active neurotoxin from the venom of Bungarus multicinctus , was radiolabelled with 125 I and its binding to synaptic membranes from rat brain was analyzed. The interaction of these binding sites with those for dendrotoxin (a convulsant polypeptide from mamba venom) and mast‐cell‐degranulating peptide (from bee venom) was examined in the light of the known effects of all three toxins on voltage‐dependent K + currents. 2 When measured in Krebs/phosphate buffer, the binding appeared monotonic at low concentrations of radioiodinated β‐bungarotoxin ( K d 0.4 nM; B max 0.42 pmol/mg protein); higher concentrations of labelled toxin revealed an additional binding component of lower affinity, but computer analysis of the data failed to provide well‐defined estimates of its K d and B max values. 3 Equilibrium binding experiments conducted in imidazole‐based buffers yielded distinctly biphasic Scatchard plots; computer analysis of the data revealed two populations of sites [ K d 0.26 (±0.30) nM and 6.14 (±5.68) nM; B max 0.16 (± 0.20) and 2.65 (± 1.21) pmol/mg protein]. 4 In Krebs medium, β‐bungarotoxin was a very weak antagonist of the binding of 125 I‐labelled dendrotoxin. In imidazole medium, however, the efficacy of the inhibition was markedly increased; analysis of this inhibition showed it to be non‐competitive. 5 Dendrotoxin inhibited the binding of radioiodinated β‐bungarotoxin in Krebs medium with high potency, although the interaction was by a complex, non‐competitive mechanism. 6 Mast‐cell‐degranulating peptide inhibited non‐competitively the binding of both radiolabelled dendrotoxin and β‐bungarotoxin but with relatively low potency. 7 A speculative schematic model of the dendrotoxin β‐bungarotoxin/mast‐cell‐degranulating peptide binding component(s) is proposed. Findings are discussed in terms of the likely involvement of these sites with voltagedependent K + ‐channel proteins.