Liu Li, Boyd Stefanie D, Bulla Lee A, Winkler Duane D
Department of Biological Sciences, The University of Texas at Dallas, Richardson, TX 75083, USA.
CustomGene, LLC, Tioga, TX 76271, USA.
J Proteomics Bioinform. 2018;11(11):201-210. doi: 10.4172/0974-276X.1000487. Epub 2018 Dec 11.
The bacterium (Bt) produces protoxin proteins in parasporal crystals. Proteolysis of the protoxin generates an active toxin which is a potent microbial insecticide. Additionally, Bt toxin genes have been introduced into genetically modified crops to produce insecticidal toxins which protect crops from insect invasion. The insecticidal activity of Cry toxins is mediated by specific interaction between toxins and their respective cellular receptors. One such toxin (Cry1Ab) exerts toxicity by first targeting the 12 ectodomain region (EC12) of the moth cadherin receptor BT-R. Binding promotes a highly regulated signaling cascade event that concludes in oncotic-like cell death. We previously determined that conserved sequence motifs near the N- and C-termini of EC12 are critical for toxin binding in insect cells. Here, we have established that Cry1Ab specifically binds to EC12 as a soluble heterodimeric complex with extremely high affinity (K = 19.5 ± 1.6 nM). Binding assays using Cry1Ab toxin and a fluorescently labeled EC12 revealed that the heterodimeric complex is highly specific in that no such formation occurs between EC12 and other Cry toxins active against beetle and mosquito. Disruption of one or both terminal sequence motifs in EC12 eliminates complex formation. Until now, comprehensive biophysical characterization of Cry1Ab recognition and binding by the BT-R receptor was unresolved. The findings presented here provide insight on the molecular determinants in the Cry family of toxins and should facilitate the assessment and advancement of their use as pesticidal agents.
苏云金芽孢杆菌（Bt）在伴孢晶体中产生原毒素蛋白。原毒素经蛋白水解后产生一种活性毒素，它是一种有效的微生物杀虫剂。此外，Bt毒素基因已被导入转基因作物中，以产生杀虫毒素，保护作物免受昆虫侵害。Cry毒素的杀虫活性是由毒素与其各自细胞受体之间的特异性相互作用介导的。一种这样的毒素（Cry1Ab）首先靶向蛾类钙黏蛋白受体BT-R的12个胞外结构域区域（EC12），从而发挥毒性作用。这种结合促进了一个高度调控的信号级联事件，最终导致类胀亡细胞死亡。我们之前确定，EC12的N端和C端附近的保守序列基序对于昆虫细胞中的毒素结合至关重要。在此，我们已确定Cry1Ab作为一种可溶性异二聚体复合物以极高的亲和力（K = 19.5 ± 1.6 nM）特异性结合到EC12上。使用Cry1Ab毒素和荧光标记的EC12进行的结合试验表明，该异二聚体复合物具有高度特异性，因为EC12与其他对甲虫和蚊子有活性的Cry毒素之间不会形成这种复合物。EC12中一个或两个末端序列基序的破坏会消除复合物的形成。到目前为止，BT-R受体对Cry1Ab识别和结合的全面生物物理特征尚未得到解决。此处呈现的研究结果为Cry毒素家族中的分子决定因素提供了见解，应该有助于评估和推进它们作为杀虫剂的应用。
