The Binding Characterization of Cry Insecticidal Proteins to the Brush Border Membrane Vesicles of Helicoverpa armigera, Spodoptera exigua, Spodoptera fitura and Agrotis ipsilon

Cry toxins produced by Bacillus thuringiensis （Bt） are effective biological insecticides against certain insect species. However, there are potential risks of the evolved resistance of insects to Cry toxin owing to decreased binding of toxins to target sites in the brush border membranes of the larva midgut. The Cry toxins with different binding sites in the larval midgut have been considered to be a good combination to deploy in delaying resistance evolution. Bioassay results demonstrated that the toxicity of different Cry toxins ranked differently for each species. The toxicity ranking was CrylAc〉CrylAb〉Cry2Ab for Helicoverpa armigera, CrylB〉CrylC〉Cry2Ab for Spodoptera exigua, and Cry2Ab〉CrylB〉 CrylC for S. litura. Only Cry2Ab was toxic to Agrotis ipsilon. Binding experiments were performed with ^125I-CrylAb, ^125I-CrylAc, ^125I-CrylB, ^125I-CrylC, ^125I-Cry2Ab and the brush border membranes vesicles （BBMV） from H. armigera, S. exigua, S. litura and A. ipsilon. The binding of CrylAb and CrylAc was shown to be saturable by incubating with increasing concentrations of H. armigera BBMV （Kd=（45.00＋2.01） nmol L-1 and （12.80＋0.18） nmol L^-1, respectively; Bmax=（54.95±l.79） ng and （55.44±0.91） ng, separately）. The binding of CrylB was shown to be saturable by incubating with increasing concentrations ofS. exigua BBMV （Kd=（23.26±1.66） nmol L^-1; Bmax=（65.37±1.87） ng）. The binding of ^125I-Cry toxins was shown to be non-saturable by incubating with increasing concentrations of S. litura and A. ipsilon BBMV. In contrast, CrylB and CrylC showed some combination with the BBMV orS. litura, and a certain amount of Cry2Ab could bind to the BBMV ofA. ipsilon. These observations suggest that a future strategy could be devised for the focused combination of specific cry genes in transgenic crops to control target pests, widen the spectrum of insecticide effectiveness and postpone insect resistance evolution.