Different domains of Bacillus thuringiensis delta-endotoxins can bind to insect midgut membrane proteins on ligand blots.
We investigated the role of the constituent domains of the CryIA(b) and CryIA(c) delta-endotoxins in binding to midgut epithelial cell membrane proteins of Spodoptera exigua and Manduca sexta on ligand blots. A collection of wild-type and CryIC-CryIA hybrid toxins was used for this purpose. As demonstrated elsewhere (R. A. de Maagd, M. S. G. Kwa, H. van der Klei, T. Yamamoto, B. Schipper, J. M. Vlak, W. J. Stiekema, and D. Bosch, Appl. Environ. Microbiol. 62:1537-1543, 1996), CryIA(b) domain III recognized a 205-kDa protein on S. exigua blots, while no specific binding by domain I or II could be detected. In contrast, on ligand blots of M. sexta proteins CryIA(b) domain II recognized a 210-kDa protein and CryIA(b) domain III recognized a 250-kDa protein. Domain III is responsible for the interaction of CryIA(c) with 120-kDa major binding proteins of both S. exigua and M. sexta. In addition, in M. sexta CryIA(c) also reacts with a 210-kDa binding protein through its domain I and/or domain II. These results show that besides domain II, domain III of delta-endotoxins plays a major role in binding to putative receptors on ligand blots. However, for S. exigua there was no clear correlation between binding of toxins on ligand blots and the in vivo toxicity of the toxins. These and previous results suggest that interactions of insect membrane proteins with both domain II and domain III can occur and that detection of these interactions depends on the type of binding assay used.
de Maagd
R A
RA
Department of Molecular Biology, DLO-Centre for Plant Breeding and Reproduction Research (CPRO-DLO), Wageningen, The Netherlands. R.A.deMaagd@CPRO.DLO.NL
van der Klei
H
H
Bakker
P L
PL
Stiekema
W J
WJ
Bosch
D
D
D000595:Amino Acid Sequence; D000818:Animals; D001413:Bacillus thuringiensis / Q000472:pathogenicity*; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000378:metabolism*; D001427:Bacterial Toxins / Q000378:metabolism*; D002352:Carrier Proteins / Q000032:analysis*; D004731:Endotoxins / Q000378:metabolism*; D006460:Hemolysin Proteins; D018402:Manduca; D008565:Membrane Proteins / Q000378:metabolism*; D008969:Molecular Sequence Data; D018411:Spodoptera
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D001427:Bacterial Toxins; D002352:Carrier Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; D008565:Membrane Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis