Publication: Targeted mutagenesis of loop residues in the receptor-binding domain of the Bacillus thuringiensis Cry4Ba toxin affects larvicidal activity
Issued Date
2005-01-15
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ISSN
03781097
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2-s2.0-11144278658
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Mahidol University
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SCOPUS
Bibliographic Citation
FEMS Microbiology Letters. Vol.242, No.2 (2005), 325-332
Suggested Citation
Tipparat Tuntitippawan, Panadda Boonserm, Gerd Katzenmeier, Chanan Angsuthanasombat Targeted mutagenesis of loop residues in the receptor-binding domain of the Bacillus thuringiensis Cry4Ba toxin affects larvicidal activity. FEMS Microbiology Letters. Vol.242, No.2 (2005), 325-332. doi:10.1016/j.femsle.2004.11.026 Retrieved from: https://repository.li.mahidol.ac.th/handle/123456789/16383
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Title
Targeted mutagenesis of loop residues in the receptor-binding domain of the Bacillus thuringiensis Cry4Ba toxin affects larvicidal activity
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Abstract
Loop residues in domain II of Bacillus thuringiensis Cry δ-endotoxins have been demonstrated to be involved in insecticidal specificity. In this study, selected residues in loops β6-β7 (S387SPS390), β8-β9 (S410, N411, T413, T415, E417and G418) and β10-β11 (D454YNS457) in domain II of the Cry4Ba mosquito-larvicidal protein were changed individually to alanine by PCR-based directed mutagenesis. All mutant toxins were expressed in Escherichia coli JM109 cells as 130-kDa protoxins at levels comparable to the wild type. Only E. coli cells that express the P389A, S410A, E417A, Y455A or N456A mutants exhibited a loss in toxicity against Aedes aegypti mosquito larvae of approximately 30% when compared to the wild type. In addition, E. coli cells expressing double mutants, S410A/E417A or Y455A/N456A, at wild-type levels revealed a significantly higher loss in larvicidal activity of approximately 70%. Similar to the wild-type protoxin, both double mutant toxins were structurally stable upon solubilisation and trypsin activation in carbonate buffer, pH 9.0. These results indicate that S410and E417in the β8-β9 loop, and Y455and N456in the β10-β11 loop are involved in larvicidal activity of the Cry4Ba toxin. © 2004 Federation of European Microbiological Societies. Published by Elsevier B.V. All rights reserved.