Tigue N J, Jacoby J, Ellar D J
Department of Biochemistry, Cambridge University, United Kingdom.
Appl Environ Microbiol. 2001 Dec;67(12):5715-20. doi: 10.1128/AEM.67.12.5715-5720.2001.
The insecticidal Cry toxins produced by the bacterium Bacillus thuringiensis are comprised of three structural domains. Domain I, a seven-helix bundle, is thought to penetrate the insect epithelial cell plasma membrane through a hairpin composed of alpha-helices 4 and 5, followed by the oligomerization of four hairpin monomers. The alpha-helix 4 has been proposed to line the lumen of the pore, whereas some residues in alpha-helix 5 have been shown to be responsible for oligomerization. Mutation of the Cry1Ac1 alpha-helix 4 amino acid Asn135 to Gln resulted in the loss of toxicity to Manduca sexta, yet binding was still observed. In this study, the equivalent mutation was made in the Cry1Ab5 toxin, and the properties of both wild-type and mutant toxin counterparts were analyzed. Both mutants appeared to bind to M. sexta membrane vesicles, but they were not able to form pores. The ability of both N135Q mutants to oligomerize was also disrupted, providing the first evidence that a residue in alpha-helix 4 can contribute to toxin oligomerization.
苏云金芽孢杆菌产生的杀虫Cry毒素由三个结构域组成。结构域I是一个七螺旋束，被认为通过由α-螺旋4和5组成的发夹结构穿透昆虫上皮细胞质膜，随后四个发夹单体发生寡聚化。有人提出α-螺旋4排列在孔的内腔，而α-螺旋5中的一些残基已被证明负责寡聚化。将Cry1Ac1的α-螺旋4氨基酸Asn135突变为Gln导致对烟草天蛾的毒性丧失，但仍观察到结合。在本研究中，在Cry1Ab5毒素中进行了等效突变，并分析了野生型和突变型毒素对应物的特性。两种突变体似乎都能与烟草天蛾膜囊泡结合，但它们无法形成孔。两种N135Q突变体的寡聚化能力也受到破坏，这首次证明α-螺旋4中的一个残基可促进毒素寡聚化。
