title: Fusion proteins containing insect-specific toxins as pest control agents: snowdrop lectin delivers fused insecticidal spider venom toxin to insect haemolymph following oral ingestion.
abstract: The mannose-specific snowdrop lectin (Galanthus nivalis agglutinin: GNA), when fed to insects, binds to the gut epithelium and passes into the haemolymph. The ability of GNA to act as a carrier protein to deliver an insecticidal spider venom neurotoxin (Segestria florentina toxin 1: SFI1) to the haemolymph of lepidopteran larvae was investigated. Constructs encoding SFI1 and an SFI1/GNA fusion protein were expressed in Pichia pastoris. The insecticidal activity of purified recombinant proteins on injection was found to be comparable to published values for SfI1 purified from spider venom [Toxicon 40 (2002) 125]. Whereas neither GNA nor SFI1 alone showed acute toxicity when fed to larvae of tomato moth (Lacanobia oleracea), feeding SFI1/GNA fusion at 2.5% of dietary proteins was insecticidal to first stadium larvae, causing 100% mortality after 6 days. The protein also showed a significant, dose dependent, toxicity towards fourth and fifth stadium larvae, with growth reduced by up to approximately 90% over a 4-day assay period compared to controls. Delivery of intact SFI1/GNA to the haemolymph in these insects was shown by western blotting; haemolymph samples from fusion-fed larvae contained a GNA-immunoreactive protein of the same molecular weight as the SFI1/GNA fusion. SFI1/GNA and similar fusion proteins offer a novel and effective approach for delivering haemolymph active toxins by oral administration, which could be used in crop protection by expression in transgenic plants.
authors.affiliation: Central Science Laboratory, Department for Environment, Food and Rural Affairs, Sand Hutton, York YO41 1LZ, UK.
authors.forename: Elaine
authors.initials: E
authors.lastname: Fitches
authors.forename: Martin G
authors.initials: MG
authors.lastname: Edwards
authors.forename: Christopher
authors.initials: C
authors.lastname: Mee
authors.forename: Eugene
authors.initials: E
authors.lastname: Grishin
authors.forename: Angharad M R
authors.initials: AM
authors.lastname: Gatehouse
authors.forename: John P
authors.initials: JP
authors.lastname: Edwards
authors.forename: John A
authors.initials: JA
authors.lastname: Gatehouse
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism; D004068:Digestive System Physiological Phenomena; D005247:Feeding Behavior; D006458:Hemolymph / Q000378:metabolism; D007306:Insecticides; D007814:Larva / Q000378:metabolism; D007915:Lepidoptera / Q000378:metabolism; D037241:Mannose-Binding Lectins / Q000378:metabolism; D009498:Neurotoxins; D037121:Plant Lectins / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D013111:Spider Venoms / Q000302:isolation & purification; D013112:Spiders / Q000737:chemistry / Q000502:physiology
chemical_list: D002352:Carrier Proteins; D007306:Insecticides; D037241:Mannose-Binding Lectins; D009498:Neurotoxins; D037121:Plant Lectins; D011993:Recombinant Fusion Proteins; D013111:Spider Venoms; C063117:snowdrop lectin