European Molecular Biology Laboratory (EMBL), Hamburg Outstation, c/o DESY, Hamburg, Germany.
FEBS Lett. 2010 Feb 19;584(4):669-74. doi: 10.1016/j.febslet.2009.12.057. Epub 2010 Jan 18.
In prokaryotes, operon encoded proteins often form protein-protein complexes. Here, we show that the native structure of operons can be used to efficiently overexpress protein complexes. This study focuses on operons from mycobacteria and the use of Mycobacterium smegmatis as an expression host. We demonstrate robust and correct stoichiometric expression of dimers to higher oligomers. The expression efficacy was found to be largely independent of the intergenic distances. The strategy was successfully extended to express mycobacterial protein complexes in Escherichia coli, showing that the operon structure of gram-positive bacteria is also functional in gram-negative bacteria. The presented strategy could become a general tool for the expression of large quantities of pure prokaryotic protein complexes for biochemical and structural studies.
在原核生物中，操纵子编码的蛋白质通常形成蛋白质-蛋白质复合物。在这里，我们表明，操纵子的天然结构可用于有效地过表达蛋白质复合物。本研究集中于分枝杆菌的操纵子，并利用耻垢分枝杆菌作为表达宿主。我们证明了二聚体到更高寡聚体的稳健和正确的化学计量表达。发现表达功效在很大程度上与基因间距离无关。该策略成功扩展到在大肠杆菌中表达分枝杆菌蛋白复合物，表明革兰氏阳性菌的操纵子结构在革兰氏阴性菌中也是功能性的。所提出的策略可以成为表达大量纯原核蛋白复合物用于生化和结构研究的通用工具。
