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.
基金
Supported by Natural Science Foundation of China(81560611)
Natural Science Foundation of Hainan Province(No.317170)
