文献类型：期刊文献

中文题名：不同拷贝数及间隔肽序列对毕赤酵母分泌表达EXA的影响

英文题名：Influence of Copy Number and Spacer Peptides on Secretive Expression of exendin-4 Analog in Pichia pastoris

作者：王晖[1];钱江潮[1];庄英萍[1];储炬[1];张嗣良[1]

机构：[1]华东理工大学生物反应器工程国家重点实验室,国家生化工程技术研究中心(上海),上海200237

年份：2008

卷号：34

期号：1

起止页码：40

中文期刊名：华东理工大学学报（自然科学版）

外文期刊名：Journal of East China University of Science and Technology

收录：CSTPCD;;EI(收录号：20081311171209);Scopus;北大核心:【北大核心2004】；CSCD:【CSCD2011_2012】；

基金：国家科技部“863”项目(2002AA217021);上海市科委二次专利开发项目(047252058)

语种：中文

中文关键词：间隔肽;exendin-4类似物;多拷贝串联;毕赤酵母;分泌表达

外文关键词：spacer peptide; exendin-4; multi-copy in tandem; pichia pastoris ; secretive expression

摘要：通过多拷贝串联表达，以及在分泌信号序列和目的蛋白N端之间插入几个氨基酸的间隔短肽，以提高exendin-4类似物串联体EXA在重组毕赤酵母中的分泌表达水平。实验结果表明：2～5个EXA串联插入均比单个EXA插入有利于提高表达水平，而4拷贝EXA串联体的表达产物较多滞留在胞内。在EXA的N端增加间隔肽可促进蛋白分泌，与不合间隔肽序列的4拷贝EXA串联表达载体相比，含有糖基化住点（NTTEEGEPK）和肠激酶识别位点（DDDDK）间隔肽的插入，使蛋白表达水平分别提高了4倍和9倍。
In order to promote the secretive expression of the analog of exendin-4 in recombinant Pichia pastoris, we adopted two strategies： one was to express the multi-copy recombinant protein in tandem and the other was to introduce spacer peptides which contain several amino acids between secretive sequence and the N terminal of the recombinant protein. Results indicated that 2-5 copies＇ EXA in tandem were better than one copy EXA for EXA expression, while most 4-EXA protein was retained in the cell. In addition, our research proved that introducing a spacer peptide at the N terminal of the EXA could increase the secretive expression level. In comparison to the vector containing 4-exa gene without spacer peptide, the expression level of vectors containing glycosylation site sequence（NTTEEGEPK） and enterokinase recognition site sequence （DDDDK） as spacer was increased by 4 and 9 times, respectively.