通过断裂内含肽介导的反式剪接合成大的蛋白

中国生物工程杂志

2009, Vol. 29

Issue (12): 74-78

技术与方法

通过断裂内含肽介导的反式剪接合成大的蛋白

张静,刘环,周晶,刘建华^**

上海交通大学生命科学技术学院上海 200240

Large Protein Production Through Split Intein-Mediated Trans-Splicing

ZHANG Jing,LIU Huan,ZHOU Jing,LIU Jian-hua

College of Life Science & Biotechnology, Shanghai Jiao Tong University, Shanghai 200240,China

全文: PDF(519 KB) HTML

摘要：

大肠杆菌难以表达大的蛋白，毒性蛋白以及膜蛋白，“Npu DnaE内含肽表达系统“使这些蛋白的表达成为可能。该系统的基本原理是：在特定位点处将目标基因（编码T7 RNA聚合酶的基因）断裂成两部分，然后分别与Npu DnaE内含肽的N端，C端片段融合，两种融合基因分别表达纯化，在体外将两种融合蛋白等摩尔比混合即可产生有功能的T7 RNA聚合酶。理论上，该体系也可用于合成其他大的蛋白，毒性蛋白或膜蛋白。

关键词： 大的蛋白; 毒性蛋白; 膜蛋白; Npu DnaE内含肽; T7 RNA聚合酶

Abstract:

Npu DnaE intein was used to produce some large proteins, which were difficult to obtain through conventional expression systems. A T7 expression system was described, by which the gene of T7 RNA polymerase is split into two pieces, and each piece fuses with Npu DnaE N and C terminal sequences respectively. Functional T7 RNA polymerase is created by mixing the two kinds of fusion constructs in vitro.The approach of split inteinmediated production of large proteins, in theory, readily generalizable to the purification of other large, cytotoxic or membrane proteins.

Key words: Large cytotoxic or membrane proteins Npu DnaE intein T7 RNA polymerase

收稿日期: 2009-09-09 出版日期: 2009-12-21

ZTFLH:

Q819

基金资助:

国家“973”计划子课题（2007CB914500）资助项目

通讯作者: 刘建华 E-mail: jianhualiudl@sjtu.edu.cn

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引用本文:

张静刘环周晶刘建华. 通过断裂内含肽介导的反式剪接合成大的蛋白[J]. 中国生物工程杂志, 2009, 29(12): 74-78.

ZHANG Jing, LIU Huan, ZHOU Jing, LIU Jian-Hua. Large Protein Production Through Split Intein-Mediated Trans-Splicing. China Biotechnology, 2009, 29(12): 74-78.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/ 或 https://manu60.magtech.com.cn/biotech/CN/Y2009/V29/I12/74

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