Role of the structurally disordered N- and C-terminal residues in the Janus-faced atracotoxins

Abstract

The Janus-faced atracotoxins (J-ACTXs) are a family of insect-specific excitatory toxins isolated from the venom of Australian funnel-web spiders (genera Atrax and Hadronyche). In addition to a classical cystine knot motif, these toxins contain a rare vicinal disulfide bond. While the vicinal disulfide is known to be critical for insecticidal activity, the role of other residues in toxin function remains to be determined. In this study, we probed the role of the structurally disordered N- and C-terminal residues using a panel of recombinant mutants of the prototypic family member J-ACTX-Hv1c. We found that the structurally disordered C-terminal residues (Glu 36 and Pro 37) were dispensable for toxin function. However, whereas deletion of Ala 1 had minimal impact on toxin function, deletion of both Ala 1 and Ile 2 decreased insecticidal activity more than 70-fold. We propose that Ile 2 forms a part of the target binding site of J-ACTX-Hv1c.