Publication: Aromaticity of tyr-202 in the α4-α5 loop is essential for toxicity of the Bacillus thuringiensis Cry4A toxin
Issued Date
2004-05-31
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12258687
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2-s2.0-3142734918
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Mahidol University
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SCOPUS
Bibliographic Citation
Journal of Biochemistry and Molecular Biology. Vol.37, No.3 (2004), 292-297
Suggested Citation
Walairat Pornwiroon, Gerd Katzenmeier, Sakol Panyim, Chanan Angsuthanasombat Aromaticity of tyr-202 in the α4-α5 loop is essential for toxicity of the Bacillus thuringiensis Cry4A toxin. Journal of Biochemistry and Molecular Biology. Vol.37, No.3 (2004), 292-297. Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/21183
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Title
Aromaticity of tyr-202 in the α4-α5 loop is essential for toxicity of the Bacillus thuringiensis Cry4A toxin
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Abstract
The current model for the mechanism of action of the Bacillus thuringiensis Cry δ-endotoxins involves the penetration of the α4-α5 hairpin into the target midgut epithelial cell membranes, followed by pore formation. In this study, PCR-based mutagenesis was employed to identify a critical residue within the α4-α5 loop of the 130-kDa Cry4A mosquito-larvicidal protein. Alanine-substitutions of two charged (Asp-198 and Asp-200) and four polar (Asn-190, Asn-195, Tyr-201 and Tyr-202) residues in the α4-α5 loop were performed. Like the wildtype, all of the mutant toxins were over-expressed as inclusion bodies in Escherichia coli. When E. coli cells expressing each mutant toxin were bioassayed against Aedes aegypti larvae, larvicidal activity was completely abolished for the substitution of only Tyr-202, while replacements at the other positions still retained a high level of toxicity. Further replacement of Tyr-202 with an aromatic side chain, phenylalanine, did not affect the toxicity. These results revealed a crucial role in toxin activity for the conserved aromatic residue at the 202 position within the α4-α5 loop of the Cry4A toxin.