Correlative microscopy reveals the nanoscale morphology of E. coli -derived supported lipid bilayers
Correlative microscopy reveals the nanoscale morphology of E. coli -derived supported lipid bilayers
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相关显微镜揭示了大肠杆菌衍生的支持脂质双层的纳米级形态
DOI：
10.1101/2021.11.04.467316
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发表时间：
2021
期刊：
影响因子：
--
通讯作者：
Bali K
中科院分区：
文献类型：
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作者：
Bali K
支撑脂质双分子层（slb）是分子生物学研究的重要工具。该领域的一项突破是利用来自细胞膜的囊泡来形成slb。这些新的支持双分子层，由天然和合成组分组成，为研究蛋白质-蛋白质相互作用、蛋白质-配体相互作用和其他脂质膜特性提供了一个生理学相关的系统。这些复杂的双层系统有希望，但尚未在其组成，天然与合成成分的比例和膜蛋白含量方面完全表征。在这里，我们描述了一种使用结构照明显微镜（SIM）的相关原子力（AFM）方法，用于精确定位由合成部分和来自大肠杆菌外膜囊泡（omv）的部分脂质组成的复杂脂质双层。我们利用SIM可以提供的增强分辨率和分子特异性来识别这些双层结构中感兴趣的区域，以及AFM提供的原子尺度分辨率来创建双层结构的详细地形图。因此，我们能够理解两种不同的脂质组分（天然的和合成的）在双层中混合的方式，量化在双层中结合的细菌膜的数量，并直接可视化这些双层与细菌特异性膜结合蛋白的相互作用。我们的工作为准确理解omv衍生slb的组成和性质奠定了基础，并建立了相关的AFM/ SIM作为表征纳米级复杂体系的方法。
Supported lipid bilayers (SLBs) made from reconstituted lipid vesicles are an important tool in molecular biology. A breakthrough in the field has come with the use of vesicles derived from cell membranes to form SLBs. These new supported bilayers, consisting both of natural and synthetic components, provide a physiologically relevant system on which to study protein-protein interactions as well as protein-ligand interactions and other lipid membrane properties. These complex bilayer systems hold promise but have not yet been fully characterised in terms of their composition, ratio of natural to synthetic component and membrane protein content. Here, we describe a method of correlative atomic force (AFM) with structured illumination microscopy (SIM) for the accurate mapping of complex lipid bilayers that consist of a synthetic fraction and a fraction of lipids derived fromEscherichia coliouter membrane vesicles (OMVs). We exploit the enhanced resolution and molecular specificity that SIM can offer to identify areas of interest in these bilayers and the atomic scale resolution that the AFM provides to create detailed topography maps of the bilayers. We are thus able to understand the way in which the two different lipid fractions (natural and synthetic) mix within the bilayers, quantify the amount of bacterial membrane incorporated in the bilayer and directly visualise the interaction of these bilayers with bacteria-specific, membrane-binding proteins. Our work sets the foundation for accurately understanding the composition and properties of OMV-derived SLBs and establishes correlative AFM/ SIM as a method for characterising complex systems at the nanoscale.