芋螺毒素MrIA的串联表达、纯化及生物活性鉴定

doi:10.13523/j.cb.20160512

中国生物工程杂志

2016, Vol. 36

Issue (5): 81-88 DOI: 10.13523/j.cb.20160512

研究报告

芋螺毒素MrIA的串联表达、纯化及生物活性鉴定

朱晓鹏^1,2,3, 张亚宁¹, 于津鹏¹, 李晓丹¹, 罗素兰^1,2, 长孙东亭^1,2

1. 海南大学热带生物资源教育部重点实验室海口市海洋药物重点实验室海口 570228;
2. 海南大学海洋学院海口 570228;
3. 海南大学园艺园林学院海口 570228

Tandem Expression, Purification and Bioactivity Characterization of Conotoxin MrIA

ZHU Xiao-peng^1,2,3, ZHANG Ya-ning¹, YU Jin-peng¹, LI Xiao-dan¹, LUO Su-lan^1,2, ZHANGSUN Dong-ting^1,2

1. Key Laboratory of Tropical Biological Resources, Ministry of Education, Key Laboratory for Marine Drugs of Haikou, Hainan University, Haikou 570228, China;
2. College of Marine Science, Hainan University, Haikou 570228, China;
3. College of Horticulture and Landscapes, Hainan University, Haikou 570228, China

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摘要：

芋螺毒素(Conotoxins,CTxs)是一类特异作用于离子通道和膜受体的小分子多肽,是研究受体结构和功能及其相关疾病的重要工具。MrIA是进入临床研究可用于镇痛治疗的一种T超家族的芋螺毒素。传统获取芋螺毒素是通过化学合成的方法,成本高、产量低。实验利用串联表达技术,构建原核表达载体,在大肠杆菌(Escherichia coli,E. coli)中成功表达了芋螺毒素MrIA(recombinant MrIA,rMrIA)。通过溴化氰切割和纯化,最终1L菌液可获得纯度高达95%的rMrIA 30mg。小鼠热板实验表明,rMrIA具有较好的镇痛活性。这为大量获得MrIA以及其他芋螺毒素小肽的表达提供了方法。

关键词： 融合表达; 热板实验; 芋螺毒素MrIA; 纯化; 电生理

Abstract:

CTxs are small peptides which specific target variety of voltage-gated ion channels and membrane receptor. Conotoxins are important tools for the study of receptors structure and function and their related diseases. MrIA is a T-superfamily conotoxin which was entering clinical research for analgesia. The traditional methods to acquire conotoxin are chemical synthesis, while these methods are cost-consuming but low yields. Tandem repeats of mria gene fragment were constructed and ligated with prokaryotic expression vector. The rMrIA was expressed successfully in E.coli. 95% purity rMrIA were obtained by CNBr cleavage and purification, 30mg rMrIA could be obtained from 1L media eventually. Mice hot plate test showed that rMrIA had good analgesic activity. An effective method to obtain large quantities of MrIA and a model for other small conotoxins expression were provided.

Key words: Conotoxin MrIA Electrophysiology Purification Fusion expression Hot plate test

收稿日期: 2015-10-09 出版日期: 2016-02-15

ZTFLH:

Q78

基金资助:

国家自然科学基金(41366002),海南省自然科学基金(314041),长江学者和创新团队发展计划(IRT_15R15)资助项目

通讯作者: 长孙东亭 E-mail: zhangsundt@163.com

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引用本文:

朱晓鹏, 张亚宁, 于津鹏, 李晓丹, 罗素兰, 长孙东亭. 芋螺毒素MrIA的串联表达、纯化及生物活性鉴定[J]. 中国生物工程杂志, 2016, 36(5): 81-88.

ZHU Xiao-peng, ZHANG Ya-ning, YU Jin-peng, LI Xiao-dan, LUO Su-lan, ZHANGSUN Dong-ting. Tandem Expression, Purification and Bioactivity Characterization of Conotoxin MrIA. China Biotechnology, 2016, 36(5): 81-88.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/10.13523/j.cb.20160512 或 https://manu60.magtech.com.cn/biotech/CN/Y2016/V36/I5/81

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