Rietveld A G, Koorengevel M C, de Kruijff B
Department of Biomembrane, Utrecht University, The Netherlands.
EMBO J. 1995 Nov 15;14(22):5506-13. doi: 10.1002/j.1460-2075.1995.tb00237.x.
The construction of a mutant Escherichia coli strain which cannot synthesize phosphatidylethanolamine provides a tool to study the involvement of non-bilayer lipids in membrane function. This strain produces phosphatidylglycerol and cardiolipin (CL) as major membrane constituents and requires millimolar concentrations of divalent cations for growth. In this strain, the lipid phase behaviour is tightly regulated by adjustment of the level of CL which favours a nonbilayer organization in the presence of specific divalent cations. We have used an in vitro system of inverted membrane vesicles to study the involvement of non-bilayer lipids in protein translocation in the secretion pathway. In this system, protein translocation is very low in the absence of divalent cations but can be enhanced by inclusion of Mg2+, Ca2+ or Sr2+ but not by Ba2+ which is unable to sustain growth of the mutant strain and cannot induce a non-bilayer phase in E. coli CL dispersions. Alternatively, translocation in cation depleted vesicles could be increased by incorporation of the non-bilayer lipid DOPE (18:1) but not by DMPE (14:0) or DOPC (18:1), both of which are bilayer lipids under physiological conditions. We conclude that non-bilayer lipids are essential for efficient protein transport across the plasma membrane of E. coli.
构建一种无法合成磷脂酰乙醇胺的突变型大肠杆菌菌株，为研究非双层脂质在膜功能中的作用提供了一种工具。该菌株产生磷脂酰甘油和心磷脂（CL）作为主要膜成分，生长需要毫摩尔浓度的二价阳离子。在该菌株中，脂质相行为通过调节CL水平来严格调控，在特定二价阳离子存在的情况下，CL有利于形成非双层结构。我们使用了一种体外反向膜泡系统来研究非双层脂质在分泌途径中蛋白质转运的作用。在这个系统中，在没有二价阳离子的情况下蛋白质转运非常低，但可以通过加入Mg2 +、Ca2 +或Sr2 +来增强，而Ba2 +则不能，因为Ba2 +无法维持突变菌株的生长，也不能在大肠杆菌CL分散体中诱导非双层相。或者，通过掺入非双层脂质DOPE（18:1）可以增加阳离子耗尽的膜泡中的转运，但DMPE（14:0）或DOPC（18:1）则不能，这两种脂质在生理条件下都是双层脂质。我们得出结论，非双层脂质对于大肠杆菌质膜上高效的蛋白质运输至关重要。
