Structure-function relationships of the major neurotoxin from the sea anemone Stichodactyla helianthus with a new sodium channel receptor site

We have determined that ShN I, a 48-residue type 2 sea anemone toxin, delays the inactivation of the Na channel in lobster olfactory somas. The receptor for ShN I was identified in vesicle preparations of neuronal tissues from both crustaceans and mammals; however, the KD values for the former is more than 1,000 fold lower for the later. The binding of [125I]-ShN I to this receptor was determined to be unaffected by Anemonia sulcata II, depolarization of the membrane, or veratridine. ShN I was unable to displace [125I]-Androctonus austrialis Hector II, whereas unlabeled AaH II and As II displaced the labeled scorpion toxin from rat brain synaptosomes. This is the first characterization of a new Na channel receptor site which specifically binds type 2 anemone toxins. To study the interactions that specific amino acid residues of ShN I have with this receptor, we developed a strategy using solid phase peptide synthesis. Prior to the synthesis of analogs to ShN I, we assembled the native ShN I sequence and reoxidized the three intramolecular disulfide bonds. Chemical, physical, and pharmacological characterization of the purified synthetic ShN I showed it to be indistinguishable from the natural toxin