Ercan Bilge, Low Wen-Yi, Liu Xuejun, Chng Shu-Sin
Department of Chemistry , National University of Singapore , Singapore 117543.
School of Pharmacy , University College London , London WC1N 1AX , United Kingdom.
Biochemistry. 2019 Jan 15;58(2):114-119. doi: 10.1021/acs.biochem.8b00897. Epub 2018 Oct 8.
The outer membrane (OM) of Gram-negative bacteria is a permeability barrier that impedes the entry of external insults, such as antibiotics and bile salts. This barrier function depends critically on the asymmetric lipid distribution across the bilayer, with lipopolysaccharides (LPS) facing outside and phospholipids (PLs) facing inside. In Escherichia coli, the OmpC-Mla system is believed to maintain OM lipid asymmetry by removing surface exposed PLs and shuttling them back to the inner membrane (IM). How proteins in the pathway interact to mediate PL transport across the periplasm is not known. Evidence for direct transfer of PLs between these proteins is also lacking. In this study, we mapped the interaction surfaces between the two PL-binding proteins, MlaC and MlaD, using site-specific in vivo photo-cross-linking, and obtained a physical picture for how these proteins may transfer PLs. Furthermore, we demonstrated using purified proteins that MlaD spontaneously transfers PLs to MlaC, suggesting that the latter has a higher affinity for PLs. Our work provides insights into the mechanism of bacterial intermembrane lipid transport important for the maintenance of OM lipid asymmetry.
革兰氏阴性菌的外膜是一种渗透屏障，可阻止外部侵害物质（如抗生素和胆汁盐）的进入。这种屏障功能关键取决于双层膜上脂质的不对称分布，其中脂多糖（LPS）面向外部，磷脂（PLs）面向内部。在大肠杆菌中，OmpC-Mla系统被认为通过去除表面暴露的PLs并将它们运回内膜（IM）来维持外膜脂质不对称性。该途径中的蛋白质如何相互作用以介导PLs跨周质转运尚不清楚。也缺乏PLs在这些蛋白质之间直接转移的证据。在本研究中，我们使用位点特异性体内光交联绘制了两种PL结合蛋白MlaC和MlaD之间的相互作用表面，并获得了这些蛋白质可能如何转移PLs的物理图像。此外，我们使用纯化蛋白证明MlaD可自发地将PLs转移至MlaC，这表明后者对PLs具有更高的亲和力。我们的工作为细菌膜间脂质转运机制提供了见解，这对于维持外膜脂质不对称性很重要。
