Selection of a δ-latroinsectotoxin fragment from Latrodectus tredecimguttatus venom for efficient bacterial expression
Keywords:
biological plant protection, heterologous expression, polyclonal antibodiesAbstract
Toxins from the venom of predatory and parasitoid arthropods, such as scorpions, spiders, and parasitoid wasps, are used as promising tools in plant protection against insect pests. In many cases, polyclonal antibodies can be created to study a single molecule in the laboratory, which allows for specific methods of detection and purification. In the present study, heterologous expression of δ-latroinsectotoxin from the venom of the black widow spider Latrodectus tredecimguttatus, as well as its N- and C- terminal fragments, was carried out in Escherichia coli cells at various temperatures and aeration conditions. Regardless of the expression conditions, the accumulation of the recombinant protein in large quantities in the bacterial cells as inclusion bodies was observed only for the C-terminal fragment of the toxin. The high efficiency of expression of this fragment allowed the use of the insoluble protein fraction of the bacterial homogenate for the immunization of mice and the production of antibodies to δ-latroinsectotoxin. The antigen-binding activity of the immunoglobulins was confirmed by immunoblotting of the synthesized toxin fragment in bacteria using a vector with removed leader sequences. The antibodies obtained can be used to detect the black widow spider toxin during its expression in various systems, such as in the creation of genetically modified entomopathogenic fungi or viruses.
