A SNAKE TOXIN AGAINST MUSCARINIC ACETYLCHOLINE-RECEPTORS - AMINO-ACID-SEQUENCE, SUBTYPE SPECIFICITY AND EFFECT ON GUINEA-PIG ILEUM

The sequence of muscarinic toxin 1 (MT1) from Dendroaspis angusticeps (green mamba) was determined (66 amino acids, M(r) 7509). The central part, peptide 25-40, is rich in hydrophobic amino acids, which is a characteristic of muscarinic toxins, MT1 started to inhibit [H-3]-NMS (N-methylscopolamine) binding to synaptosomal membranes of porcine brain (contains all five receptor subtypes) at about 1 nM and to membranes from pig heart muscle (only subtype m2) at about 1 mu M. Binding of [H-3]-AF-DX 384 to heart was inhibited with an IC50 of 14 mu M and to brain in two steps. In the first step (IC50 = 32 nM) binding decreased by 37%, indicating that the toxin acted on mi or m4 receptors, each accounting for about 40% of total receptor content. The second step was similar to the effect on heart. Pirenzepine inhibited binding of [I-125]-MT1 to brain receptors with an IC50 of 6.5 nM, corresponding to a K-i of about 6 nM. Literature values of K-i for pirenzepine are 16-18 nM for mi and greater than or equal to 120 mM for other subtypes, This indicates binding to mi receptors, [I-125]- MT1 bound to brain with a K-d of 20 nM and a Hill coefficient of 1.0, i.e. one toxin molecule per receptor. In guinea-pig ileum, MT1 (670 nM) produced a rapid contraction, reversible by atropine. The toxin may be an agonist, but might also cause contraction by inducing acetylcholine release by a different mechanism.