Isolation, synthesis and pharmacological characterization of δ‐palutoxins IT, novel insecticidal toxins from the spider Paracoelotes luctuosus (Amaurobiidae)

Abstract

Four novel insecticidal toxins were isolated from the venom of the spider Paracoelotes luctuosus (Araneae: Amaurobiidae) and named d-palutoxins IT1 to IT4. The four toxins are homologous 36±37 amino acid peptides reticulated by four disulfide bridges and three have amidated C-terminal residues. The d-palutoxins are highly homologous with the previously described m-agatoxins and curtatoxins (77±97%). The four peptides demonstrated significant toxicity against larvae of the crop pest Spodoptera litura (Lepidoptera: Noctuidae) in a microinjection bioassay, with LD 50 values in the 9±50 mg per g of insect range. This level of toxicity is equivalent to that of several of the most active scorpion toxins used in the development of recombinant baculoviruses, and the d-palutoxins appear to be insect specific. Electrophysiological experiments demonstrated that d-palutoxin IT1, the most active toxin acts by affecting insect sodium channel inactivation, resulting in the appearance of a late-maintained sodium current, in a similar fashion to insecticidal scorpion a and a-like toxins and is thus likely to bind to channel receptor site 3. However, d-palutoxin IT1 was distinguished by its lack of effect on peak sodium conductance, on the early phase of sodium current inactivation and the absence of a shift in the activation voltage of the sodium channels. d-Palutoxins are thus proposed as new insecticidal toxins related to the a and a-like scorpion toxins. They will be useful both in the development of recombinant baculoviruses in agrochemical applications and also as molecular probes for the investigation of molecular mechanisms of insect selectivity and structure and function of sodium channels.Keywords: baculovirus; insecticidal toxins; scorpion toxins; sodium channel; spider toxins.Intensive use of synthetic chemicals as the sole strategy for insect control is being reconsidered as public tolerance of the negative effects of synthetic pesticides increases and pressure mounts against widespread and indiscriminate use of agrochemicals. Microbial insecticides, and in particular the group of insect-specific baculoviruses [1], have been proposed as an attractive alternative in integrated pest management programs or specific applications. In the past, substantial research efforts have been devoted to the development of recombinant baculoviruses expressing proteins deleterious to insects with the goal of producing selective, fast-acting pathogens to be used in combination with other pest-control agents [2,3]. One of the most successful approaches to increase the speed of kill in baculoviruses has been genetic engineering into the viruses of arthropod toxin genes with neurotoxic effects on the insect nervous system. Significant decreases in the time of the appearance of lethal effects and the reduction of crop damage have been obtained with recombinant viruses expressing toxins [1,4,5]. The AaIT scorpion toxin from Androctonus australis and other scorpion toxins affecting sodium channel kinetics such as LqhaIT from Leiurus quinques...

Enzymatic, Antioxidant, Antimicrobial, and Insecticidal Activities of Pleurotus pulmonarius and Pycnoporus cinnabarinus Grown Separately in an Airlift Reactor

“…Insecticidal activities of CEs were evaluated by the larvae microinjection technique (Corzo et al 2000). Diatraea magnifactella larvae were injected in the pronotum with a volume of CE equivalent to 10% of their weight using glass capillary pipettes, and they were placed in plastic 55-mm Petri dishes with artificial Meridic diet.…”

Peptide toxins that selectively target insect NaVand CaVchannels

“…A quite different mode of action is seen with the δ-palutoxins (δ-PaluITs), a family of 36-37-residue peptide toxins from the spider Paracoelotes luctuosus that selectively modulate the activity of insect Na V channels. 89 Using an isolated cockroach axon preparation and cloned Drosophila Para/TipE Na V channels expressed in Xenopus oocytes, the δ-PaluITs have been shown to slow insect Na V channel inactivation with no shift in the voltage dependence of activation. They also have no effect on vertebrate Na V 1.2 channels at concentrations up to 10 μM.…”

SPider Neurotoxins Targeting Voltage-Gated Sodium Channels

“…Interestingly, the toxin failed to modify TTX-resistant Nav channels in rat DRG neurons but showed a 30-fold higher potency for inhibition of cardiac TTX-resistant INa than for insect or mammalian neuronal Nav channels, thus discriminating cardiac from neuronal Nav channel isoforms. are a family of four 36-37 residue peptides that selectively modulate insect Nav channels (Corzo et al, 2000) (Fig. 2C).…”