An alternative method of enhancing the expression level of heterologous protein in Escherichia coli
An alternative method of enhancing the expression level of heterologous protein in Escherichia coli
复制标题
增强大肠杆菌中异源蛋白表达水平的另一种方法
DOI：
10.1016/j.bbrc.2014.10.149
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发表时间：
2014
影响因子：
3.1
通讯作者：
Yao Wenbing
中科院分区：
文献类型：
--
作者：
Yin Jun;Tian Hong;Bao Lichen;Dai Xin;Gao Xiangdong;Yao Wenbing
虽然有许多策略选择可以在大肠杆菌中实现高水平的基因表达，但并不是每个基因都能在这个生物体中高效表达。通过研究大肠杆菌翻译起始区(TIR)的二级结构与基因表达的关系，我们建立了一种简单的方法来设计合适的TIR序列(从−35到+36)，以满足我们所需要的特定表达水平。用该方法过表达人体液坏死因子α和HER2/neu胞外区蛋白(aa23-146)。大肠菌群均已实现。表达差异的出现主要与翻译起始效率和mRNA二级结构的稳定性有关，因为实时荧光RT-PCR分析的细胞内mRNA水平非常相似。我们的方法可以克服翻译启动的空间障碍，从而促进翻译的顺利进行，获得外源蛋白的高表达。
Though numerous strategy options are available for achieving high expression levels of genes inEscherichia coli, not every gene can be efficiently expressed in this organism. By investigating the relationship between the mRNA secondary structure of translational initiation region (TIR) and gene expression inE.coli, we establish a simple method to design sequences of appropriate TIR (from −35 to +36) that meet a specific expression level as we need. Using this method, overexpression of native human humor necrosis factor α and extracellular domain of Her2/neu protein (aa 23–146) inE. coliwere achieved. Differences in expression appeared was mainly related to the efficiency of translation initiation and the stability of mRNA secondary structure, because the intracellular mRNA levels analyzed by real-time RT-PCR were quite similar. Our approach can overcome the steric hindrance of translation startup, and therefore promote translation smoothly to acquire high expression of exogenous protein.