Comparative Pharmacology And Cloning Of Two Novel Sodium Channels

Scorpion toxins have been found lacking effect on gating of scorpion's own sodiumchannel, but the molecular mechanism involved in the insensitivity remains a mystery.In this study, first, the binding properties of pharmacologically distinct scorpion toxinson scorpion (Buthus martensii) and its closely relative spider (Ornithoctonus huwena)nerve synaptosomes was investigated using the Biomolecular Interaction Analysis.Results show that the affinity of scorpion nerve synaptosomes for these toxins ismarkedly decreased and the decrease in binding is mainly derived from an increaseddissociation rate, contrasting with the "quasi-irreversibility" of the binding in spidersynaptosomes. Comparing their relative affinities suggests the insensitivity towardsscorpion toxins is confined solely to scorpion sodium channel, but not a character ofall arachnid ones. Second, the sodium channel cDNA from these two species wascloned and their deduced amino acid sequences were compared for conservation andnatural mutation through evolution. Surprisingly, scorpion sodium channel is found topossess two extra residues (Leu-Pro) in the outer end of the IIS4 when compared toother arachnid or vertebrate counterparts. Third, mutant of rNav1.2a comprising thetwo extra residues at the corresponding region is found insensitive to β-like toxinBmK AS, contrasting with a negative shift of the voltage dependence of activation ofwild-type rNav1.2a. Combined, the results explore the molecular evidence forscorpion's adaptive evolution to its own sodium channel toxins. Meanwhile, the twonovel scorpion and spider sodium channel proteins may serve as good templates tostudy the interaction of polypeptide neurotoxins with sodium channels. Furthermore,comparative analysis based on different sensitivity of scorpion and spider sodiumchannels towards sea anemone toxin as well as sequence similarity between scorpionand sea anemone toxins suggests a possible common origin of scorpion and seaanemone sodium channel toxins. The evolution route of diverse scorpion toxins is alsodiscussed through constructing a reliable phylogenetic tree, taking into account theworldwide distribution of toxins and the zoogeographic dispersion of the studiedgenera.