Restoration of the crystallization of altered delta-endotoxins Cry1Ac, by the promotion of their in vivo integration into the Bacillus thuringiensis native crystals.
Cry1Ac is one of the most-studied Bacillus thuringiensisdelta-endotoxins. Structurally, it is divided into two domains: the N-terminal half corresponding to the toxic component and the C-terminal half corresponding to the region responsible for the crystal formation. We engineered Cry1Ac delta-endotoxins modified in their N-terminal part and studied the effect of such modifications on crystallization and delta-endotoxin production. When expressed in an acrystalliferous B. thuringiensis strain, Cry1Ac(*) and Cry1AcDelta, variants with four point mutations and a deletion, respectively, could not form crystals. However, when expressed in a crystalliferous strain, these altered proteins were shown to interact with the endogenous delta-endotoxins and cocrystallize with them, forming atypical crystals observed by electron microscopy. This cocrystallization of the altered delta-endotoxins with the endogenous ones led to a decrease in delta-endotoxin production (27%) by the corresponding recombinant B. thuringiensis strains. This ability of altered delta-endotoxins containing an intact C-terminal part to cocrystallize with native ones could be exploited to promote the crystallization of foreign proteins by fusing them with the C-terminal part of Cry1A delta-endotoxins.
Dammak
Mariam
M
Laboratory of Biopesticides, Centre of Biotechnology of Sfax, Sfax, Tunisia.
Tounsi
Slim
S
Hamadou
Dorra Ben
DB
Abdelkafi
Lobna
L
Schultz
Patrick
P
Jaoua
Samir
S
D001413:Bacillus thuringiensis / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D003460:Crystallization; D004731:Endotoxins / Q000235:genetics / Q000378:metabolism*; D006460:Hemolysin Proteins / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D046529:Microscopy, Electron, Transmission; D017354:Point Mutation; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion
D000083722:Bacillus thuringiensis Toxins; D001426:Bacterial Proteins; D004731:Endotoxins; D006460:Hemolysin Proteins; D011994:Recombinant Proteins; C030956:insecticidal crystal protein, Bacillus Thuringiensis