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Scorpion insect neurotoxin LqhIT2 is a promising oral biopesticide: high‐level preparation in Pichia pastoris and bioactivity assays
Pest Management Science ( IF 3.8 ) Pub Date : 2024-12-26 , DOI: 10.1002/ps.8626
Hongbo Li, Cheng Tian, Jing Chen, Yuanxian Xia
Pest Management Science ( IF 3.8 ) Pub Date : 2024-12-26 , DOI: 10.1002/ps.8626
Hongbo Li, Cheng Tian, Jing Chen, Yuanxian Xia
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BACKGROUNDDiscovering insecticidal proteins with high activity and strict insect specificity and applying them to the biological control of insect pests is of great significance. Oral LqhIT2 has insecticidal activity, which most other insecticidal neurotoxin proteins do not have, but the large‐scale preparation of the toxin is difficult and one of the obstacles to determining its anti‐insect potential for biological control.RESULTSIn this study, the expression level of recombinant LqhIT2 (rLqhIT2) in Pichia pastoris was as high as 1.4 g/L under high‐cell‐density fermentation conditions. A purified rLqhIT2 at a final concentration of 0.5 ng/mL inhibited the proliferation of insect Sf9 cells, with typical insect cell oncosis observed, while 200 ng/mL rLqhIT2 was non‐toxic to mammalian cells. The injection of silkworms caused the typical depressive symptoms of flaccid paralysis and death. At a dose of 1 μg/g body weight, 46.7% of the silkworms died within 48 h after injection, and their mortality rate was related to the injection dose. Feeding purified rLqhIT2 to fifth instar Pieris rapae larvae demonstrated its oral insecticidal activity, with a mortality rate 66.7% after 3 days of feeding at a concentration of 10 μg/cm2 cabbage leaf.CONCLUSIONThe rLqhIT2 has high insect cells specificity and insecticidal activity when injected and or delivered orally, and may have better application prospects than other reported insect neurotoxins. We describe a method for large‐scale production of functional rLqhIT2 by fed‐batch fermentation using Pichia pastoris , which will further facilitate its application in biological control of agricultural insect pests. © 2024 Society of Chemical Industry.
中文翻译:
蝎子昆虫神经毒素 LqhIT2 是一种很有前途的口服生物农药:毕赤酵母和生物活性测定中的高水平制备
背景 发现具有高活性和严格昆虫特异性的杀虫蛋白并将其应用于害虫的生物防治具有重要意义。口服 LqhIT2 具有大多数其他杀虫神经毒素蛋白所不具备的杀虫活性,但毒素的大规模制备很困难,并且是确定其生物防治抗虫潜力的障碍之一。结果本研究在高细胞密度发酵条件下,毕赤酵母中重组 LqhIT2 (rLqhIT2) 的表达水平高达 1.4 g/L。终浓度为 0.5 ng/mL 的纯化 rLqhIT2 抑制昆虫 Sf9 细胞的增殖,观察到典型的昆虫细胞癌变,而 200 ng/mL rLqhIT2 对哺乳动物细胞无毒。注射蚕引起了弛缓性麻痹和死亡的典型抑郁症状。在 1 μg/g 体重剂量下,46.7% 的蚕在注射后 48 h 内死亡,其死亡率与注射剂量有关。将纯化的 rLqhIT2 喂食给 5 龄 Pieris rapae 幼虫表现出其口服杀虫活性,以 10 μg/cm2 的浓度饲喂 3 天后死亡率为 66.7%。结论rLqhIT2 在注射和/或口服给药时具有较高的昆虫细胞特异性和杀虫活性,可能比其他已报道的昆虫神经毒素具有更好的应用前景。我们描述了一种使用 Pichia pastoris 通过补料分批发酵大规模生产功能性 rLqhIT2 的方法,这将进一步促进其在农业害虫生物防治中的应用。© 2024 化工学会.
更新日期:2024-12-26
中文翻译:
![](https://scdn.x-mol.com/jcss/images/paperTranslation.png)
蝎子昆虫神经毒素 LqhIT2 是一种很有前途的口服生物农药:毕赤酵母和生物活性测定中的高水平制备
背景 发现具有高活性和严格昆虫特异性的杀虫蛋白并将其应用于害虫的生物防治具有重要意义。口服 LqhIT2 具有大多数其他杀虫神经毒素蛋白所不具备的杀虫活性,但毒素的大规模制备很困难,并且是确定其生物防治抗虫潜力的障碍之一。结果本研究在高细胞密度发酵条件下,毕赤酵母中重组 LqhIT2 (rLqhIT2) 的表达水平高达 1.4 g/L。终浓度为 0.5 ng/mL 的纯化 rLqhIT2 抑制昆虫 Sf9 细胞的增殖,观察到典型的昆虫细胞癌变,而 200 ng/mL rLqhIT2 对哺乳动物细胞无毒。注射蚕引起了弛缓性麻痹和死亡的典型抑郁症状。在 1 μg/g 体重剂量下,46.7% 的蚕在注射后 48 h 内死亡,其死亡率与注射剂量有关。将纯化的 rLqhIT2 喂食给 5 龄 Pieris rapae 幼虫表现出其口服杀虫活性,以 10 μg/cm2 的浓度饲喂 3 天后死亡率为 66.7%。结论rLqhIT2 在注射和/或口服给药时具有较高的昆虫细胞特异性和杀虫活性,可能比其他已报道的昆虫神经毒素具有更好的应用前景。我们描述了一种使用 Pichia pastoris 通过补料分批发酵大规模生产功能性 rLqhIT2 的方法,这将进一步促进其在农业害虫生物防治中的应用。© 2024 化工学会.