BIOTINYLATION OF BACILLUS-THURINGIENSIS INSECTICIDAL CRYSTAL PROTEINS

Biotinylation of Bacillus thuringiensis insecticidal crystal proteins (ICPs) was evaluated for its potential use in an alternative ICP scree ning method and in the characterization of ICP receptors. In vivo biol ogical activity of CryIA(b), as inferred from bioassays with Manduca s exta and Ostrinia nubilalis and from histopathological effects on O. n ubilalis midgut cells induced by force feeding, was not affected by bi otinylation at moderate biotinylation ratios. A competitive radiorecep tor assay showed that there was only a minor reduction in binding affi nity of biotin-labeled CryIA(b) for M. sexta brush border membrane ves icles. On midgut tissue sections, the binding pattern along the midgut epithelium and the staining intensity of biotinylated ICPs detected w ith streptavidin-enzyme conjugate were virtually identical to the bind ing pattern and staining intensity of native CryIA(b) detected with an tibodies. The specificity of biotinylated ICP binding to larval midgut tissue was demonstrated by performing homologous competition experime nts. The relationship between different ICP receptor types in Plutella xylostella, as inferred from radioligand binding studies, was confirm ed by the results of heterologous competition experiments performed wi th biotinylated and native ICPs.