Escherichia coli fusion carrier proteins act as solubilizing agents for recombinant uncoupling protein 1 through interactions with GroEL
Escherichia coli fusion carrier proteins act as solubilizing agents for recombinant uncoupling protein 1 through interactions with GroEL
复制标题
DOI：
10.1016/j.bbrc.2005.05.164
复制
发表时间：
2005-08-05
影响因子：
3.1
通讯作者：
Sluse, FE
中科院分区：
文献类型：
--
作者：
Douette, P;Navet, R;Sluse, FE
将重组蛋白融合到高可溶性配偶体上经常用于防止重组蛋白在大肠杆菌中聚集。此外，原核分子伴侣的共过表达可以增加正确折叠的重组蛋白的量。为了了解融合蛋白的溶解性增强，我们设计了两种重组蛋白，其由解偶联蛋白1（UCP 1）（一种线粒体膜蛋白）与MBP或NusA融合组成。我们能够表达MBP-UCP 1和NusA-UCP 1的可溶形式，尽管UCP 1具有高疏水性。此外，可溶性融合蛋白的产量取决于催化多肽折叠的GroEL的共过表达。MBP-UCP 1以与GroEL的非共价复合物的形式表达。纯化MBP-UCP 1/GroEL，并通过动态光散射、凝胶过滤和电子显微镜进行表征。我们的研究结果表明，MBP和NusA作为增溶剂，迫使重组蛋白通过细菌伴侣途径的背景下融合蛋白。(c)2005年爱思唯尔公司All rights reserved.
Fusing recombinant proteins to highly soluble partners is frequently used to prevent aggregation of recombinant proteins in Escherichia coli. Moreover, co-overexpression of prokaryotic chaperones can increase the amount of properly folded recombinant proteins. To understand the solubility enhancement of fusion proteins, we designed two recombinant proteins composed of uncoupling protein 1 (UCP1), a mitochondrial membrane protein, in fusion with MBP or NusA. We were able to express soluble forms of MBP-UCP1 and NusA-UCP1 despite the high hydrophobicity of UCP1. Furthermore, the yield of soluble fusion proteins depended on co-overexpression of GroEL that catalyzes folding of polypeptides. MBP-UCP1 was expressed in the form of a non-covalent complex with GroEL. MBP-UCP1/GroEL was purified and characterized by dynamic light scattering, gel filtration, and electron microscopy. Our findings suggest that MBP and NusA act as solubilizing agents by forcing the recombinant protein to pass through the bacterial chaperone pathway in the context of fusion protein. (c) 2005 Elsevier Inc. All rights reserved.