Expression and purification of Huwentoxin-I in baculovirus system
Expression and purification of Huwentoxin-I in baculovirus system
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DOI：
10.1016/j.pep.2005.02.012
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发表时间：
2005-06-01
影响因子：
1.6
通讯作者：
An, CC
中科院分区：
文献类型：
--
作者：
Ji, WJ;Zhang, XY;An, CC
虎纹捕鸟蛛毒素-I（HWTX-I）是从虎纹捕鸟蛛（Orinithoctonus huwena）毒液中分离得到的一种新的神经毒素。基于其生物活性，HWTX-1可开发为临床用途的止痛药。通过细菌或酵母表达系统生产HWTX-1导致产量低，纯化的蛋白被证明具有比天然蛋白低的生物活性。因此，我们首次采用杆状病毒表达系统在Sf 9细胞中表达HWTX-I基因。重组HWTX-I通过Western印迹识别，然后在天然条件下通过镍螯合亲和层析纯化。重组HWTX-Ⅰ的氨基酸序列与天然型完全相同，对肌肉传递的作用与天然型相似。这些结果表明杆状病毒表达系统和天然纯化策略是生产活性HWTX-1的可行方法。(c)2005年爱思唯尔公司All rights reserved.
Huwentoxin-I (HWTX-I) is a novel neurotoxin isolated from the venom of Orinithoctonus huwena. Based on its biological activity, HWTX-I could be developed as a pain-killer for clinical purpose. Production of HWTX-I by the bacterium or yeast expression systems resulted in poor yields and the purified protein was proved to have lower biological activity than that of native one. So, for the first time, we introduced a new method to express HWTX-I gene in Sf9 cells using baculovirus expression system. Recombinant HWTX-I was recognized by Western blotting and then purified by nickel-chelating affinity chromatography under native conditions. Recombinant HWTX-I showed identical amino acid sequence as native form and exhibited similar effect on muscular transmission with that of native form. These results indicate that the baculovirus expression system and native purification strategy are viable ways to produce active HWTX-I. (c) 2005 Elsevier Inc. All rights reserved.