Isolated domain II and III from the Bacillus thuringiensis Cry1Ab delta-endotoxin binds to lepidopteran midgut membranes.
The DNA fragment encoding Cry1Ab domain II-III (45.3 kDa) was cloned and expressed. Domain II-III is expressed in low yields. In vitro binding analysis to Manduca sexta and Trichoplusia ni larval midgut tissue sections demonstrated that domain II-III fragment bound along the microvilli of the midgut epithelium, indicating that this fragment retains binding functionality in the absence of domain I. Binding of domain II-III to the midgut brush border membrane proteins from T. ni larvae indicated that Cry1Ab toxin and domain II-III bind to the same 150 kDa protein. In contrast, in M. sexta membranes, Cry1Ab toxin binds to 200 and 120 kDa proteins, and domain II-III only binds to the 200 kDa protein. Finally, binding assays with isolated brush border membrane vesicles showed that the interaction of domain II-III with the membrane vesicles is highly reversible, supporting the proposition that the integration of domain I into the membrane could participate in the irreversible binding of the toxin. These studies confirm that this part of the toxin is involved in binding interactions and could be separated as a discrete fragment that conserves at least part of its functionality.
Flores
H
H
Instituto de Biotecnologia, Universidad Nacional Autónoma de México, Cuernavaca, Mor.
Soberón
X
X
Sánchez
J
J
Bravo
A
A
D000818:Animals; D001413:Bacillus thuringiensis; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism*; D001427:Bacterial Toxins; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D017931:DNA Primers; D004064:Digestive System / Q000378:metabolism*; D004731:Endotoxins / Q000737:chemistry* / Q000378:metabolism*; D004926:Escherichia coli; D006460:Hemolysin Proteins; D007150:Immunohistochemistry; D007814:Larva; D007915:Lepidoptera; D018402:Manduca; D008871:Microvilli / Q000378:metabolism*; D008961:Models, Structural; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010572:Pest Control, Biological; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D017931:DNA Primers; D004731:Endotoxins; D006460:Hemolysin Proteins; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis