A Novel Family of Insect-Selective Peptide Neurotoxins Targeting Insect Large-Conductance Calcium-Activated K(+) Channels Isolated from the Venom of the Theraphosid Spider Eucratoscelus constrictus

Amer Soc Pharmacology Experimental Therapeutics
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Journal Article
Molecular Pharmacology, 2011, 80 (1), pp. 1 - 13
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Spider venoms are actively being investigated as sources of novel insecticidal agents for biopesticide engineering. After screening 37 theraphosid spider venoms, a family of three new "short-loop" inhibitory cystine knot insecticidal toxins (kappa-TRTX-Ec2a, kappa-TRTX-Ec2b, and kappa-TRTX-Ec2c) were isolated and characterized from the venom of the African tarantula Eucratoscelus constrictus. Whole-cell patch-clamp recordings from cockroach dorsal unpaired median neurons revealed that, despite significant sequence homology with other theraphosid toxins, these 29-residue peptides lacked activity on insect voltage-activated sodium and calcium channels. It is noteworthy that kappa-TRTX-Ec2 toxins were all found to be high-affinity blockers of insect large-conductance calcium-activated K+ (BKCa) channel currents with IC50 values of 3 to 25 nM. In addition, kappa-TRTX-Ec2a caused the inhibition of insect delayed-rectifier K+ currents, but only at significantly higher concentrations. kappa-TRTX-Ec2a and kappa-TRTX-Ec2b demonstrated insect-selective effects, whereas the homologous kappa-TRTX-Ec2c also resulted in neurotoxic signs in mice when injected intracerebroventricularly. Unlike other theraphosid toxins, kappa-TRTX-Ec2 toxins induce a voltage-independent channel block, and therefore, we propose that these toxins interact with the turret and/or loop region of the external entrance to the channel and do not project deeply into the pore of the channel. Furthermore, kappa-TRTX-Ec2a and kappa-TRTX-Ec2b differ from other theraphotoxins at the C terminus and positions 5 to 6, suggesting that these regions of the peptide contribute to the phyla selectivity and are involved in targeting BKCa channels. This study therefore establishes these toxins as tools for studying the role of BKCa channels in insects and lead compounds for the development of novel insecticides.
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