School of Biomedical Sciences, University of Nottingham, Nottingham, United Kingdom.
PLoS One. 2012;7(6):e38677. doi: 10.1371/journal.pone.0038677. Epub 2012 Jun 7.
Lipopolysaccharide (LPS) is a major constituent of bacterial outer membranes where it makes up the bulk of the outer leaflet and plays a key role as determinant of bacterial interactions with the host. Membrane-free LPS is known to activate T-lymphocytes through interactions with Toll-like receptor 4 via multiprotein complexes. In the present study, we investigate the role of cholesterol and membrane heterogeneities as facilitators of receptor-independent LPS binding and insertion, which underpin bacterial interactions with the host in symbiosis, pathogenesis and cell invasion. We use fluorescence spectroscopy to investigate the interactions of membrane-free LPS from intestinal gram-negative organisms with cholesterol-containing model membranes and with T-lymphocytes. LPS preparations from Klebsiella pneumoniae and Salmonella enterica were found to bind preferentially to mixed lipid membranes by comparison to pure PC bilayers. The same was observed for LPS from the symbiote Escherichia coli but with an order of magnitude higher dissociation constant. Insertion of LPS into model membranes confirmed the preference for sphingomyelin/cholesterol-containing systems. LPS insertion into Jurkat T-lymphocyte membranes reveals that they have a significantly greater LPS-binding capacity by comparison to methyl-β-cyclodextrin cholesterol-depleted lymphocyte membranes, albeit at slightly lower binding rates.
脂多糖（LPS）是细菌外膜的主要组成部分，构成外膜叶的大部分，作为决定细菌与宿主相互作用的关键因素发挥作用。已知无膜脂多糖通过与 Toll 样受体 4 相互作用，通过多蛋白复合物激活 T 淋巴细胞。在本研究中，我们研究了胆固醇和膜异质性作为促进受体非依赖性 LPS 结合和插入的因素的作用，这些因素是细菌在共生、发病机制和细胞入侵中与宿主相互作用的基础。我们使用荧光光谱法研究了来自肠道革兰氏阴性生物的无膜 LPS 与含胆固醇的模型膜以及与 T 淋巴细胞的相互作用。与纯 PC 双层相比，来自肺炎克雷伯菌和沙门氏菌的 LPS 制剂被发现优先结合混合脂质膜。共生菌大肠杆菌的 LPS 也是如此，但解离常数高一个数量级。通过将 LPS 插入模型膜来确认对鞘磷脂/胆固醇含量系统的偏好。LPS 插入 Jurkat T 淋巴细胞膜表明，与甲基-β-环糊精胆固醇耗尽的淋巴细胞膜相比，它们具有更大的 LPS 结合能力，尽管结合速率略低。
