Preparation of Conotoxin MrVIB by Genetic Engineering Technology  

作　　者：Weiwei GUAN Jie HOU Xia ZHONG Na WEI Junqing ZHANG Bingmiao GAO 

机构地区：[1]Hainan Provincial Key Laboratory of Research and Development of Tropical Medicinal Plants, School of Pharmacy, Hainan Medical University, Haikou 571199,China

出　　处：《Agricultural Biotechnology》2017年第4期28-31,37,共5页农业生物技术（英文版）

基　　金：Supported by Natural Science Foundation of China(81560611);Natural Science Foundation of Hainan Province(No.317170)

摘　　要：[ Objective] The disulfide-rich conotoxin MrV1B was produced by simple and fast genetic engineering method, to find new efficient ways for the synthesis of natural active conotoxins. [Method] Primers of conotoxin gene MrVIB were synthesized to construct expression vectors pET22b（ ＋ ）/His-Xa-MrVIB and pET32a/Trx-EK-MrV1B, which were transformed into BL21 （DE3）pLysS and expressed under induction by IPTG. Recombinant proteins were purified by affinity chromatography using Ni-NTA agarose column, and the expression of the recombinant proteins was analyzed by Tricine-SDS-PAGE electrophoresis. [ Result] The recombinant conotoxins His-Xa-MrVIB and Trx-EK-MrVIB were effectively expressed in E. coli, and purified by one-step affinity chromatography, and the purity of the recombinant conotoxins was greater than 90%. [ Conclusion] The conotoxin MrVIB was effectively secreted and expressed by genetic engineering method, which could solve the problems in chemical synthesis of conotoxins including low yield, high cost and difficult purification.[ Objective] The disulfide-rich conotoxin MrV1B was produced by simple and fast genetic engineering method, to find new efficient ways for the synthesis of natural active conotoxins. [Method] Primers of conotoxin gene MrVIB were synthesized to construct expression vectors pET22b（ ＋ ）/His-Xa-MrVIB and pET32a/Trx-EK-MrV1B, which were transformed into BL21 （DE3）pLysS and expressed under induction by IPTG. Recombinant proteins were purified by affinity chromatography using Ni-NTA agarose column, and the expression of the recombinant proteins was analyzed by Tricine-SDS-PAGE electrophoresis. [ Result] The recombinant conotoxins His-Xa-MrVIB and Trx-EK-MrVIB were effectively expressed in E. coli, and purified by one-step affinity chromatography, and the purity of the recombinant conotoxins was greater than 90%. [ Conclusion] The conotoxin MrVIB was effectively secreted and expressed by genetic engineering method, which could solve the problems in chemical synthesis of conotoxins including low yield, high cost and difficult purification.

关 键 词：CONOTOXINS Escherichia coli Genetic Engineering Recombinant Expression Separation and Purification 

分 类 号：Q78[生物学—分子生物学] TQ464.9[化学工程—制药化工]