Bacteria co-transformed with recombinant proteins and chaperones cloned in independent plasmids are suitable for expression tuning
Bacteria co-transformed with recombinant proteins and chaperones cloned in independent plasmids are suitable for expression tuning
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DOI：
10.1016/j.jbiotec.2003.10.025
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发表时间：
2004-04-08
影响因子：
4.1
通讯作者：
De Marco, V
中科院分区：
文献类型：
--
作者：
de Marco, A;De Marco, V
由于代谢限制，通常会阻止几种重组蛋白在同一细菌细胞中的有效过度表达。然而，共同产生大量复合物亚基或表达大量伴侣和折叠酶的可能性在技术上可能非常有用。我们开发了一个基于三个载体的系统。其中两个受 IPTG 调控，能够重组表达六种分子伴侣，第三个是阿拉伯糖诱导型，包含目标蛋白的序列。以这种方式，伴侣和靶蛋白的独立诱导和表达水平是可能的。数据显示，通过在细胞中引入更多质粒，可以防止 pET 载体的表达泄漏，并且重组蛋白竞争其表达。事实上，其中一种重组蛋白的高诱导率可以关闭其他重组蛋白的积累。第一个信息用于最大化有毒蛋白的表达，同时利用重组蛋白之间的交叉抑制来调节和优化靶蛋白表达并诱导分子伴侣辅助的聚集靶蛋白体内重新折叠。 (C) 2004 Elsevier B.V. 保留所有权利。
The efficient over-expression of several recombinant proteins in the same bacterial cell is usually prevented due to metabolic limitations. Nevertheless, the possibility to co-produce high amounts of the sub-units of a complex or to express a wide set of chaperones and foldases could be technologically very useful.We developed a system based on three vectors. Two are under IPTG regulation and enable the recombinant expression of six chaperones, the third one is arabinose-inducible and harbours the sequence for the target protein. In such a way the independent induction and the level of expression of both chaperones and target protein is possible. The data show that the expression leakage from pET vectors was prevented by the introduction of further plasmids in the cell and that the recombinant proteins compete for their expression. In fact, the high rate induction of one of them could switch off the accumulation of the other recombinant proteins. The first information was used to maximise the expression of toxic proteins while the cross-inhibition among recombinant proteins was exploited to modulate and optimise the target protein expression and to induce the chaperone-assisted in vivo re-folding of aggregated target protein. (C) 2004 Elsevier B.V. All rights reserved.