Nanoscale Features of Tunable Bacterial Outer Membrane Models Revealed by Correlative Microscopy.
Nanoscale Features of Tunable Bacterial Outer Membrane Models Revealed by Correlative Microscopy.
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DOI：
10.1021/acs.langmuir.2c00628
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发表时间：
2022-07-26
期刊：
影响因子：
3.9
通讯作者：
Mela, Ioanna
中科院分区：
文献类型：
--
作者：
Bali, Karan;Mohamed, Zeinab;Scheeder, Anna;Pappa, Anna-Maria;Daniel, Susan;Kaminski, Clemens F.;Owens, Roisin M.;Mela, Ioanna
抗生素耐药性的增加是一个日益严重的全球人类健康问题，具有重大的社会经济影响。了解细菌膜上发生的相互作用对于新抗生素的产生至关重要。由重构脂质囊泡制成的支撑脂质双层（SLB）已被用于模拟这些膜，但它们的实用性受到这些系统的简单性质的限制。该领域的一个突破是使用来源于革兰氏阴性细菌的外膜囊泡形成SLB，从而提供了一个更生理相关的系统。这些复杂的双层系统有希望，但尚未完全表征其组成，天然合成成分的比例，和膜蛋白含量。在这里，我们使用相关原子力显微镜（AFM）和结构照明显微镜（SIM）来准确绘制复杂脂质双层，该双层由合成部分和源自大肠杆菌外膜囊泡（OMV）的脂质部分组成。我们利用SIM可以提供的高分辨率和分子特异性来识别这些双层中的感兴趣区域，以及AFM提供的增强分辨率来创建双层的详细形貌图。因此，我们能够理解两种不同的脂质组分（天然和合成）在双层中混合的方式，并且我们可以量化并入双层中的细菌膜的量。我们证明了系统的可调谐性，通过产生双层使用OMV工程包含一个绿色荧光蛋白（GFP）结合纳米抗体与孔蛋白OmpA融合。我们能够直接可视化GFP和纳米抗体复合物之间的蛋白质-蛋白质相互作用。我们的工作为准确理解OMV衍生的SLB的组成和性质奠定了基础，以产生一个高分辨率的平台，用于研究细菌膜相互作用，以开发下一代抗生素。
The rise of antibiotic resistance is a growing worldwide human health issue, with major socioeconomic implications. An understanding of the interactions occurring at the bacterial membrane is crucial for the generation of new antibiotics. Supported lipid bilayers (SLBs) made from reconstituted lipid vesicles have been used to mimic these membranes, but their utility has been restricted by the simplistic nature of these systems. A breakthrough in the field has come with the use of outer membrane vesicles derived from Gram-negative bacteria to form SLBs, thus providing a more physiologically relevant system. These complex bilayer systems hold promise but have not yet been fully characterized in terms of their composition, ratio of natural to synthetic components, and membrane protein content. Here, we use correlative atomic force microscopy (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 from Escherichia coli outer membrane vesicles (OMVs). We exploit the high resolution and molecular specificity that SIM can offer to identify areas of interest in these bilayers and the enhanced resolution that 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, and we can quantify the amount of bacterial membrane incorporated into the bilayer. We prove the system’s tunability by generating bilayers made using OMVs engineered to contain a green fluorescent protein (GFP) binding nanobody fused with the porin OmpA. We are able to directly visualize protein–protein interactions between GFP and the nanobody complex. Our work sets the foundation for accurately understanding the composition and properties of OMV-derived SLBs to generate a high-resolution platform for investigating bacterial membrane interactions for the development of next-generation antibiotics.
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影响因子：
3.3
作者：
Piggot, Thomas J.;Holdbrook, Daniel A.;Khalid, Syma
通讯作者：
Khalid, Syma
影响因子：
6.4
作者：
Wendel S;Fischer EC;Martínez V;Seppälä S;Nørholm MH
通讯作者：
Nørholm MH
影响因子：
16.6
作者：
Lin, Yi-Chih;Chipot, Christophe;Scheuring, Simon
通讯作者：
Scheuring, Simon
DOI：
10.1007/978-1-4939-8591-3_8
发表时间：
2018-01-01
期刊：
NANOSCALE IMAGING: METHODS AND PROTOCOLS
影响因子：
--
作者：
Lv, Zhengjian;Banerjee, Siddhartha;Lyubchenko, Yuri L.
通讯作者：
Lyubchenko, Yuri L.
影响因子：
17.1
作者：
Pappa, Anna-Maria;Liu, Han-Yuan;Owens, Roisin M.
通讯作者：
Owens, Roisin M.