Restriction of intramolecular movements within the Cry1Aa toxin molecule of Bacillus thuringiensis through disulfide bond engineering |
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Authors: | Jean-Louis Schwartz Marc Juteau Pawel Grochulski Miroslaw Cygler Gabrielle Prfontaine Roland Brousseau Luke Masson |
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Institution: | Jean-Louis Schwartz, Marc Juteau, Pawel Grochulski, Miroslaw Cygler, Gabrielle Préfontaine, Roland Brousseau,Luke Masson |
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Abstract: | Disulfide bridges were introduced into Cry1Aa, a Bacillus thuringiensis lepidopteran toxin, to stabilize different protein domains including domain I α-helical regions thought to be involved in membrane integration and permeation. Bridged mutants could not form functional ion channels in lipid bilayers in the oxidized state, but upon reduction with β-mercaptoethanol, regained parental toxin channel activity. Our results show that unfolding of the protein around a hinge region linking domain I and II is a necessary step for pore formation. They also suggest that membrane insertion of the hydrophobic hairpin made of α-helices 4 and 5 in domain I plays a critical role in the formation of a functional pore. |
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Keywords: | disulfide bond |
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