Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Reexamination Certificate
1999-12-10
2002-09-24
Kaufman, Claire M. (Department: 1646)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
C435S007200, C435S069100, C435S320100, C435S325000, C435S252300, C435S254110, C435S007100, C435S006120, C530S300000, C530S350000, C536S023100, C536S023500, C536S023710
Reexamination Certificate
active
06455266
ABSTRACT:
TECHNICAL FIELD
The invention relates to receptors that bind toxins from
Bacillus thuringiensis
and thus to pesticides and pest resistance. More particularly, the invention concerns recombinantly produced receptors that bind BT toxin and to their use in assays for improved pesticides, as well as in mediation of cell and tissue destruction, dissociation, dispersion, cell-to-cell association, and changes in morphology.
BACKGROUND ART
It has long been recognized that the bacterium
Bacillus thuringiensis
(BT) produces bactericidal proteins that are toxic to a limited range of insects, mostly in the orders Lepidoptera, Coleoptera and Diptera. Advantage has been taken of these toxins in controlling pests, mostly by applying bacteria to plants or transforming plants themselves so that they generate the toxins by virtue of their transgenic character. The toxins themselves are glycoprotein products of the cry gene as described by Höfte, H. et al.
Microbiol Rev
(1989) 53:242. It has been established that the toxins function in the brush border of the insect midgut epithelial cells as described by Gill, S. S. et al.
Annu Rev Entomol
(1992)37:615. Specific binding of BT toxins to midgut brush border membrane vesicles has been reported by Hofmann, C. et al.
Proc Natl Acad Sci USA
(1988) 85:7844; Van Rie, J. et al.
Eur J Biochem
(1989) 186:239; and Van Rie, J. et al.
Appl Environ Microbiol
(1990) 56:1378.
Presumably, the toxins generated by BT exert their effects by some kind of interaction with receptors in the midgut. The purification of a particular receptor from
Manduca sexta
was reported by the present inventors in an article by Vadlamudi, R. K. et al.
J Biol Chem
(1993) 268:12334. In this report, the receptor protein was isolated by immunoprecipitating toxin-binding protein complexes with toxin-specific antisera and separating the complexes by SDS-PAGE followed by electroelution. However, to date, there has been no structural information concerning any insect receptor which binds BT toxin, nor have, to applicants' knowledge, any genes encoding these receptors been recovered.
DISCLOSURE OF THE INVENTION
The present invention is based, in part, on the isolation and characterization of a receptor that is bound by members of the BT-toxin family of insecticidal proteins, hereinafter the BT-R
1
protein. The present invention is further based on the isolation and characterization of a nucleic acid molecule that encodes the BT-toxin receptor, hereinafter BT-R
1
gene. Based on these observations, the present invention provides compositions and methods for use in identifying agents that bind to the BT-R
1
protein as a means for identifying insecticidal agent and for identifying other members of the BT-R
1
family of proteins.
REFERENCES:
patent: 4675285 (1987-06-01), Clark et al.
patent: 5071654 (1991-12-01), English
Oddou et al., Immunolgoically unrelated Heliothis sp. and Spodoptera sp. midgut membrane-proteins bindBacillus thruingeinesisCrylA (b) delta-endotoxin, Eur. J. Biochem. 212: 145-150, 1993.*
Gill, S. S. et al. (1992) “The Mode of Action ofBacillus thuringiensisEndotoxins” Ammu Rev Entomol 37:615-636.
Hofte, H. et al., (1989) “Insecticidal Crystal Proteins ofBacillus thuringiensis” Microbiol Review 53:242-255.
Hofmann, C. et al. (1988) “Specificity ofBacillus thuringiensis. . . ” Proc. Natl Acad Sci USA 85:7844-7848.
Ishihara, T. et al. (1991) “Molecular cloning and expression of a cDNA encoding the secretin receptor.” EMBO J. 10(7):1635-1641.
Lee, M.K. et al. (1992) “Location of a Bombyx mori receptor binding region on aBacillus thuringiensisdelta-Endotoxins.” J. Biol. Chem. 267(5):3315-3121.
Vadlamudi, R.K. et al. (1993) “A Specific Binding Protein from Manduca Sexta for the Insecticidal Toxin ofBacillus thuringiensisSubsp.” J. Biol Chem 268(17):12334-12340.
Vadlamudi et al. (1995) Journal of Biological Chemistry 270(10): 5490-5494.
Van Rie, J. et al. (1989) “Specificity ofBacillus thuringiensis. . . ” Eur J. Biochem 186:239-247.
Van Rie, J. et al. (1990) “Receptors on the Brush Border Membrane of the Insect Midgut as Determinants of the Specificity ofBacillus thuringiensis” Belta-Endotoxins Appl Environ Microbiol 56(5):1378-1385.
Kaufman Claire M.
Morrison & Foerster / LLP
University of Wyoming
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