Ji Wenjie, Zhang Xinyue, Hu Huicong, Chen Jiyuan, Gao Yin, Liang Songping, An Chengcai
National Laboratory of Protein Engineering and Plant Genetic Engineering, College of Life Sciences, Peking University, Beijing 100871, People's Republic of China.
Protein Expr Purif. 2005 Jun;41(2):454-8. doi: 10.1016/j.pep.2005.02.012.
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.
虎纹毒素-I（HWTX-I）是从虎纹捕鸟蛛毒液中分离出的一种新型神经毒素。基于其生物活性，HWTX-I有望开发成为一种临床用止痛药。利用细菌或酵母表达系统生产HWTX-I产量较低，且纯化后的蛋白经证实其生物活性低于天然蛋白。因此，我们首次引入了一种新方法，利用杆状病毒表达系统在Sf9细胞中表达HWTX-I基因。重组HWTX-I经蛋白质免疫印迹法鉴定后，在天然条件下通过镍螯合亲和层析进行纯化。重组HWTX-I与天然形式具有相同的氨基酸序列，并且在肌肉传递方面表现出与天然形式相似的作用。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-I的可行方法。
