Interacting Domains of Anti-Insect Scorpion Toxins and their Sodium Channel Binding Sites: Structure, Cooperative Interactions with Agrochemicals, and Application

Integrated pest management in modern crop protection may combine chemical and biological insecticides, particularly due to the risks to the environment and livestock arising from the massive use of non-selective chemicals. Thus, there is a need for safer alternatives, which target insects more specifically. Scorpions produce anti-insect selective polypeptide toxins that are biodegradable and non-toxic to warm-blooded animals. Therefore, integration of these substances into insect pest control strategies is of major importance. Moreover, clarification of the molecular basis of this selectivity may provide valuable information pertinent to their receptor sites and to the future design of peptidomimetic anti-insect specific substances. These toxins may also be important for reducing the current overuse of chemical insecticides if they produce a synergistic effect with conventional pesticides. Based on these considerations, our major objectives were: 1) To elucidate the three-dimensional structure and toxic-site of scorpion excitatory, "depressant, and anti-insect alpha toxins. 2) To obtain an initial view to the sodium channel recognition sites of the above toxins by generating peptide decoys through a phage display system. 3) To investigate the synergism between toxins and chemical insecticides. Our approach was to develop a suitable expression system for toxin production in a recombinant form and for elucidation of toxin bioactive sites via mutagenesis. In parallel, the mode of action and synergistic effects of scorpion insecticidal toxins with pyrethroids were studied at the sodium channel level using electrophysiological methods. Objective 1 was achieved for the alpha toxin, LqhaIT Zilberberg et al., 1996, 1997; Tugarinov et al., 1997; Froy et al., 2002), and the excitatory toxin, Bj-xtrIT (Oren et al., 1998; Froy et al., 1999; unpublished data). The bioactive surface of the depressant toxin, LqhIT2, has been clarified and a crystal of the toxin is now being analyzed (unpublished). Objective 2 was not successful thus far as no phages that recognize the toxins were obtained. We therefore initiated recently an alternative approach, which is introduction of mutations into recombinant channels and creation of channel chimeras. Objective 3 was undertaken at Riverside and the results demonstrated synergism between LqhaIT or AaIT and pyrethroids (Lee et al., 2002). Furthermore, negative cross-resistance between pyrethroids and scorpion toxins (LqhaIT and AaIT) was demonstrated at the molecular level. Although our study did not yield a product, it paves the way for future design of selective pesticides by capitalizing on the natural competence of scorpion toxins to distinguish between sodium channels of insects and vertebrates. We also show that future application of anti-insect toxins may enable to decrease the amounts of chemical pesticides due to their synergism.