Helix α-3 inter-molecular salt bridges and conformational changes are essential for toxicity of Bacillus thuringiensis 3D-Cry toxin family

• Published in: Scientific reports Vol. 8; no. 1; pp. 10331 - 11
• Main Authors: Pacheco, Sabino, Gómez, Isabel, Sánchez, Jorge, García-Gómez, Blanca-Ines, Czajkowsky, Daniel M., Zhang, Jie, Soberón, Mario, Bravo, Alejandra
• Format: Journal Article
• Language: English
• Published: London Nature Publishing Group UK 09-07-2018; Nature Publishing Group
• Subjects: 38/23; 38/70; 38/77; 631/337/470; 631/45/612; 631/61/252; 82/80; Bacillus thuringiensis; Crystal structure; Humanities and Social Sciences; Midgut; Monomers; multidisciplinary; Mutation; Oligomerization; Phenotypes; Salts; Science; Science (multidisciplinary); Site-directed mutagenesis; Toxicity; Toxins
• ISSN: 2045-2322; 2045-2322
• DOI: 10.1038/s41598-018-28753-8

Bacillus thuringiensis
insecticidal Cry toxins break down larval midgut-cells after forming pores. The 3D-structures of Cry4Ba and Cry5Ba revealed a trimeric-oligomer after cleavage of helices α-1 and α-2a, where helix α-3 is extended and made contacts with adjacent monomers. Molecular dynamic simulations of Cry1Ab-oligomer model based on Cry4Ba-coordinates showed that E101 forms a salt-bridge with R99 from neighbor monomer. An additional salt bridge was identified in the trimeric-Cry5Ba, located at the extended helix α-3 in the region corresponding to the α-2b and α-3 loop. Both salt-bridges were analyzed by site directed mutagenesis. Single-point mutations in the Lepidoptera-specific Cry1Ab and Cry1Fa toxins were affected in toxicity, while reversed double-point mutant partially recovered the phenotype, consistent with a critical role of these salt-bridges. The single-point mutations in the salt-bridge at the extended helix α-3 of the nematicidal Cry5Ba were also non-toxic. The incorporation of this additional salt bridge into the nontoxic Cry1Ab-R99E mutant partially restored oligomerization and toxicity, supporting that the loop between α-2b and α-3 forms part of an extended helix α-3 upon oligomerization of Cry1 toxins. Overall, these results highlight the role in toxicity of salt-bridge formation between helices α-3 of adjacent monomers supporting a conformational change in helix α-3.