Gomez S, Mateus A C, Hernandez J, Zimmermann B H
Corporacion Colombiana de Investigacion Agropecuaria, CORPOICA, Km 14 Via Mosquera, Santafe de Bogota, Colombia.
Biochem Biophys Res Commun. 2000 Dec 20;279(2):653-6. doi: 10.1006/bbrc.2000.3998.
The Andean potato weevil, Premnotrypes vorax, an insect of the order Coleoptera, is a major cause of damage to potato crops in the Andean regions of South America. The insecticidal Cry proteins from Bacillus thuringiensis are useful biological pesticides, and some are toxic to Coleopteran insects. We overexpressed recombinant, histidine-tagged Cry3Aa protein in Escherichia coli host cells. The recombinant protein was solubilized at high pH with urea, purified using Ni(2+)-nitrilo-triacetic acid affinity resin, and dialysed to lower pH and remove urea. Bioassays were performed with an insect media whose surface was spread with 70 microgram/mL purified native or recombinant toxins. First instar larvae exposed to toxin treated media for 5 days exhibited mortalities from 57% (native Cry3Aa) to 52% (recombinant Cry3Aa). Purified native and recombinant Cry3Aa proteins appeared to be equally toxic to the Andean potato weevil.
安第斯马铃薯象甲（Premnotrypes vorax），鞘翅目昆虫，是南美洲安第斯地区马铃薯作物遭受损害的主要原因。来自苏云金芽孢杆菌的杀虫Cry蛋白是有用的生物杀虫剂，其中一些对鞘翅目昆虫有毒。我们在大肠杆菌宿主细胞中过表达了重组的、带有组氨酸标签的Cry3Aa蛋白。重组蛋白在高pH值下用尿素溶解，使用镍（2+）-次氮基三乙酸亲和树脂纯化，并透析至较低pH值以去除尿素。用一种昆虫培养基进行生物测定，其表面铺有70微克/毫升纯化的天然或重组毒素。暴露于毒素处理培养基5天的一龄幼虫死亡率从57%（天然Cry3Aa）到52%（重组Cry3Aa）。纯化的天然和重组Cry3Aa蛋白对安第斯马铃薯象甲似乎具有同等毒性。
