Departamento de Genética/ERI BioTecMed, Universitat de València, Burjassot, 46100 València, Spain.
Department of Plant Protection, College of Agriculture and Natural Resources, University of Tehran, Karaj 31578-77871, Alborz, Iran.
Toxins (Basel). 2020 Feb 21;12(2):133. doi: 10.3390/toxins12020133.
(Bt) produces insecticidal proteins that are either secreted during the vegetative growth phase or accumulated in the crystal inclusions (Cry proteins) in the stationary phase. Cry1I proteins share the three domain (3D) structure typical of crystal proteins but are secreted to the media early in the stationary growth phase. In the generally accepted mode of action of 3D Cry proteins (sequential binding model), the formation of an oligomer (tetramer) has been described as a major step, necessary for pore formation and subsequent toxicity. To know if this could be extended to Cry1I proteins, the formation of Cry1Ia oligomers was studied by Western blot, after the incubation of trypsin activated Cry1Ia with insect brush border membrane vesicles (BBMV) or insect cultured cells, using Cry1Ab as control. Our results showed that Cry1Ia oligomers were observed only after incubation with susceptible coleopteran BBMV, but not following incubation with susceptible lepidopteran BBMV or non-susceptible Sf21 insect cells, while Cry1Ab oligomers were persistently detected after incubation with all insect tissues tested, regardless of its host susceptibility. The data suggested oligomerization may not necessarily be a requirement for the toxicity of Cry1I proteins.
苏云金芽孢杆菌产生的杀虫蛋白，要么在营养生长阶段分泌，要么在静止期积累在包含体（Cry 蛋白）中。Cry1I 蛋白具有晶体蛋白的典型三结构域（3D）结构，但在静止生长阶段早期分泌到培养基中。在 3D Cry 蛋白公认的作用模式（顺序结合模型）中，已描述形成寡聚体（四聚体）是一个主要步骤，对于孔形成和随后的毒性是必需的。为了知道这是否可以扩展到 Cry1I 蛋白，在用 Cry1Ab 作为对照，用胰蛋白酶激活的 Cry1Ia 孵育昆虫刷状缘膜囊泡（BBMV）或昆虫培养细胞后，通过 Western blot 研究了 Cry1Ia 寡聚体的形成。我们的结果表明，仅在与敏感鞘翅目昆虫 BBMV 孵育后观察到 Cry1Ia 寡聚体，但在与敏感鳞翅目昆虫 BBMV 或非敏感 Sf21 昆虫细胞孵育后没有观察到，而 Cry1Ab 寡聚体在与所有测试的昆虫组织孵育后持续检测到，而不管其宿主的敏感性如何。数据表明寡聚化不一定是 Cry1I 蛋白毒性的必要条件。
