title: Study of the irreversible binding of Bacillus thuringiensis Cry1Aa to brush border membrane vesicles from Bombyx mori midgut.
abstract: The binding of Bacillus thuringiensis delta-endotoxin to brush border membrane vesicles (BBMVs) from the target insect larval midgut comprises with not only a reversible but also an irreversible component. The irreversible binding of delta-endotoxin is thought to be a pathologically important factor. Here, we studied the irreversible binding of Cry1Aa to the BBMVs of Bombyx mori. The (125)I-labeled Cry1Aa bound to the solubilized brush border membrane (BBM) through rapid dissociation only, unlike the binding to BBMVs, indicating that the toxin bound to the solubilized BBM through only a reversible process. Low-temperature sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis revealed that the toxin bound irreversibly to BBMVs formed an oligomer of 220kDa, whereas that bound reversibly to the solubilized BBM did not oligomeraize. When the (125)I-labeled Cry1Aa bound irreversibly to the BBMVs was digested by proteinase K, approximately 40% of the toxin observed to be resistant to proteinase K. The molecular mass of the toxin resistant to proteinase K was 60kDa, suggesting that the irreversible binding comprise two forms. These results support the notion that the irreversible binding of the toxin to BBMVs is due to the insertion of the toxin into the lipid bilayers and oligomerization to form channels.
authors.affiliation: Department of Biological Science, Graduate School of Science, Osaka Prefecture University, 1-1 Gakuen-cho, Sakai, Osaka 599-8531, Japan. ihara@b.s.osakafu-u.ac.jp
authors.forename: Hideshi
authors.initials: H
authors.lastname: Ihara
authors.forename: Michio
authors.initials: M
authors.lastname: Himeno
mesh_terms: D000818:Animals; D001413:Bacillus thuringiensis; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D012831:Bombyx / Q000378:metabolism / Q000382:microbiology*; D002462:Cell Membrane / Q000378:metabolism / Q000382:microbiology; D004591:Electrophoresis, Polyacrylamide Gel; D004731:Endotoxins / Q000737:chemistry / Q000378:metabolism*; D041981:Gastrointestinal Tract / Q000382:microbiology; D006460:Hemolysin Proteins / Q000737:chemistry / Q000378:metabolism*; D054884:Host-Pathogen Interactions / Q000502:physiology*; D008871:Microvilli / Q000378:metabolism* / Q000382:microbiology*; D010572:Pest Control, Biological; D011485:Protein Binding
chemical_list: D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis