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Applied and Environmental Microbiology, January 2001, p. 323-329, Vol. 67, No. 1
0099-2240/01/$04.00+0 DOI: 10.1128/AEM.67.1.323-329.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
Importance of Cry1
-Endotoxin Domain II Loops
for Binding Specificity in Heliothis virescens
(L.)
Juan Luis
Jurat-Fuentes1 and
Michael J.
Adang1,2,*
Department of
Entomology1 and Department of
Biochemistry and Molecular Biology,2
University of Georgia, Athens, Georgia 30602-2603
Received 13 April 2000/Accepted 2 November 2000
We constructed a model for Bacillus
thuringiensis Cry1 toxin binding to midgut membrane
vesicles from Heliothis virescens. Brush border membrane
vesicle binding assays were performed with five Cry1 toxins that share
homologies in domain II loops. Cry1Ab, Cry1Ac, Cry1Ja, and Cry1Fa
competed with 125I-Cry1Aa, evidence that each toxin binds
to the Cry1Aa binding site in H. virescens. Cry1Ac
competed with high affinity (competition constant
[Kcom] = 1.1 nM) for 125I-Cry1Ab
binding sites. Cry1Aa, Cry1Fa, and Cry1Ja also competed for
125I-Cry1Ab binding sites, though the
Kcom values ranged from 179 to 304 nM. Cry1Ab
competed for 125I-Cry1Ac binding sites
(Kcom = 73.6 nM) with higher affinity than Cry1Aa, Cry1Fa, or Cry1Ja. Neither Cry1Ea nor Cry2Aa competed with any
of the 125I-Cry1A toxins. Ligand blots prepared from
membrane vesicles were probed with Cry1 toxins to expand the model of
Cry1 receptors in H. virescens. Three Cry1A
toxins, Cry1Fa, and Cry1Ja recognized 170- and 110-kDa proteins that
are probably aminopeptidases. Cry1Ab and Cry1Ac, and to some extent
Cry1Fa, also recognized a 130-kDa molecule. Our vesicle binding and
ligand blotting results support a determinant role for domain II loops
in Cry toxin specificity for H. virescens. The
shared binding properties for these Cry1 toxins correlate with observed
cross-resistance in H. virescens.
*
Corresponding author. Mailing address: Department of
Entomology, University of Georgia, Biosciences Building, 125 Cedar St., Athens, GA 30602-2603. Phone: (706) 542-2436. Fax: (706) 542-2640. E-mail: adang{at}arches.uga.edu.
Applied and Environmental Microbiology, January 2001, p. 323-329, Vol. 67, No. 1
0099-2240/01/$04.00+0 DOI: 10.1128/AEM.67.1.323-329.2001
Copyright © 2001, American Society for Microbiology. All rights reserved.
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