College of Horticulture, Jinling Institute of Technology, 210038 Nanjing, PR China.
Jiangsu Key Laboratory for Food Quality and Safety-State Key Laboratory Cultivation Base, Ministry of Science and Technology, Jiangsu Academy of Agricultural Sciences, 210014 Nanjing, PR China.
Int J Biol Macromol. 2019 Sep 15;137:562-567. doi: 10.1016/j.ijbiomac.2019.06.164. Epub 2019 Jun 22.
Cry2A is widely used in transgenic crops in combination with Cry1A toxins. The sensitive and robust detection of Cry2A toxin in food and the environment is necessary to monitor the safety of biopesticides. Here, we describe an approach that involves the use of phage-displayed peptide for the detection of Cry2Ad2-3-the main area of Cry2Ad2 insecticidal activity. After four rounds of panning, six positive monoclonal phage particles were obtained. Pep5 with a sequence of ACSYNHNSKCGGG displayed low cross-reactivity with other Cry toxins. The working range of detection for Cry2Ad2-3 toxin standards in the brush border membrane vesicle (BBMV)-peptide sandwich ELISA was 10-50.625 ng mL and the detection limit (LOD) was 8 ng mL. Molecular insight into the interaction of pep5 with Cry2Ad2-3 was gleaned using homology modeling and docking. Molecular docking results showed that high-affinity peptide tended to dock in the groove between the two domains of Cry2Ad2-3. The interactions within the toxin-pep5 complex were due to hydrogen bond and hydrophobic interaction. Pep5 also lead us to trap the binding region. Therefore, peptides may be a cost-efficient alternative for detecting Cry toxins and studying their mechanisms.
Cry2A 与 Cry1A 毒素结合广泛应用于转基因作物。为了监测生物农药的安全性，有必要在食品和环境中对 Cry2A 毒素进行灵敏而稳健的检测。在这里，我们描述了一种使用噬菌体展示肽检测 Cry2Ad2-3 的方法，Cry2Ad2-3 是 Cry2Ad2 杀虫活性的主要区域。经过四轮淘选，获得了六个阳性单克隆噬菌体颗粒。具有 ACSYNHNSKCGGG 序列的 Pep5 与其他 Cry 毒素的交叉反应性较低。Cry2Ad2-3 毒素标准在刷状缘膜囊泡（BBMV）-肽夹心 ELISA 中的检测范围为 10-50.625ng/mL，检测限（LOD）为 8ng/mL。使用同源建模和对接技术深入了解 pep5 与 Cry2Ad2-3 的相互作用。分子对接结果表明，高亲和力肽倾向于与 Cry2Ad2-3 的两个结构域之间的凹槽结合。毒素-肽复合物内的相互作用归因于氢键和疏水相互作用。Pep5 还使我们能够捕获结合区域。因此，肽可能是检测 Cry 毒素和研究其机制的一种具有成本效益的替代方法。
