Restoration of the crystallization of altered δ-endotoxins Cry1Ac, by the promotion of their in vivo integration into the Bacillus thuringiensis native crystals

Abstract

Cry1Ac is one of the most-studied Bacillus thuringiensisδ-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 δ-endotoxins modified in their N-terminal part and studied the effect of such modifications on crystallization and δ-endotoxin production. When expressed in an acrystalliferous B. thuringiensis strain, Cry1Ac^* and Cry1AcΔ, 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 δ-endotoxins and cocrystallize with them, forming atypical crystals observed by electron microscopy. This cocrystallization of the altered δ-endotoxins with the endogenous ones led to a decrease in δ-endotoxin production (27%) by the corresponding recombinant B. thuringiensis strains. This ability of altered δ-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 δ-endotoxins.

Bacillus thuringiensis, δ-endotoxins, cry gene, C-terminal portion, cocrystallization