Recombinant expression and purification of spider toxin, JZTX-51 and JZTX-26, from Chilobrachys jingzhao.

Jie Shao; Jiao Pan; Fang QU; Zihao Liu; Yiying Ding; Sha Luo; Xuewen Zhang; Jinjun Chen

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Recombinant expression and purification of spider toxin, JZTX-51 and JZTX-26, from Chilobrachys jingzhao.

Author: Jie SHAO ¹ ; Jiao PAN ¹ ; Fang QU ¹ ; Zihao LIU ¹ ; Yiying DING ¹ ; Sha LUO ¹ ; Xuewen ZHANG ¹ ; Jinjun CHEN ¹
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1. College of Bioscience and Biotechnology, Hunan Agricultural University, Changsha 410128, Hunan, China.
Publication Type:Journal Article
Keywords: DRG cell; fusion protein; prokaryotic expression technology; sodium channel; spider toxin
From:Chinese Journal of Biotechnology 2018;34(10):1668-1678
CountryChina
Language:Chinese
Abstract: To establish a simple, quick and effective method to get a large amount of spider toxin JZTX-26 (35 aa) and JZTX-51 (27 aa) with 3 disulfide bonds each, the mature peptides coding gene fragments were constructed and fused with maltose-binding protein (MBP) tag in an Escherichia coli expression vector pMAL-p2x. The recombinant constructs pMAL-jz26 and pMAL-jz51 were transformed and cultured in E. coli TB1 and BL21 (DE3). After being induced by isopropyl-β-d-thiogalactoside (IPTG), the periplasmic proteins were purified by amylose affinity chromatography and analyzed by SDS-PAGE. The fusion proteins were digested with factor X, and purified by Sizes-Exclusion chromatography and Reversed Phase HPLC. Molecular weights of the purified peptides were obtained by using a MALDI-TOF-TOF mass spectrometer, which were consistent with the theoretical molecular weights. Five milligram of target protein could be purified from 1 L of culture medium. The results indicate that the peptides with three disulfide bonds can be expressed by using the prokaryotic expression system with MBP tag. Our findings suggest the possibility of genetic engineering to obtain large amount of spider peptide toxins.