利用细菌外膜囊泡工程化平面革兰氏阴性外膜模拟物。
Engineering Planar Gram-Negative Outer Membrane Mimics Using Bacterial Outer Membrane Vesicles.
机构信息
Department of Chemistry, Lehigh University, Bethlehem, Pennsylvania 18015, United States.
Department of Chemical and Biomolecular Engineering, Lehigh University, Bethlehem, Pennsylvania 18015, United States.
出版信息
Langmuir. 2024 Nov 5;40(44):23289-23300. doi: 10.1021/acs.langmuir.4c02632. Epub 2024 Oct 25.
Antibiotic resistance is a major challenge in modern medicine. The unique double membrane structure of Gram-negative bacteria limits the efficacy of many existing antibiotics and adds complexity to antibiotic development by limiting transport of antibiotics to the bacterial cytosol. New methods to mimic this barrier would enable high-throughput studies for antibiotic development. In this study, we introduce an innovative approach to modify outer membrane vesicles (OMVs) from , to generate planar supported lipid bilayer membranes. Our method first involves the incorporation of synthetic lipids into OMVs using a rapid freeze-thaw technique to form outer membrane hybrid vesicles (OM-Hybrids). Subsequently, these OM-Hybrids can spontaneously rupture when in contact with SiO surfaces to form a planar outer membrane supported bilayer (OM-SB). We assessed the formation of OM-Hybrids using dynamic light scattering and a fluorescence quenching assay. To analyze the formation of OM-SBs from OM-Hybrids we used quartz crystal microbalance with dissipation monitoring (QCM-D) and fluorescence recovery after photobleaching (FRAP). Additionally, we conducted assays to detect surface-associated DNA and proteins on OM-SBs. The interaction of an antimicrobial peptide, polymyxin B, with the OM-SBs was also assessed. These findings emphasize the capability of our platform to produce planar surfaces of bacterial outer membranes, which in turn, could function as a valuable tool for streamlining the development of antibiotics.
抗生素耐药性是现代医学面临的主要挑战。革兰氏阴性菌独特的双层膜结构限制了许多现有抗生素的疗效,并通过限制抗生素向细菌细胞质的转运,使抗生素的开发变得更加复杂。模拟这种屏障的新方法将能够为抗生素的开发提供高通量研究。在这项研究中,我们介绍了一种从 中修饰外膜囊泡 (OMV) 的创新方法,以生成平面支撑脂质双层膜。我们的方法首先涉及使用快速冷冻-解冻技术将合成脂质掺入 OMV 中,以形成外膜混合囊泡 (OM-Hybrids)。随后,当这些 OM-Hybrids 与 SiO 表面接触时,它们会自发破裂,形成平面外膜支撑双层 (OM-SB)。我们使用动态光散射和荧光猝灭测定法评估了 OM-Hybrids 的形成。为了分析 OM-Hybrids 从 OM-SB 的形成,我们使用了石英晶体微天平与耗散监测 (QCM-D) 和光漂白后荧光恢复 (FRAP)。此外,我们还进行了检测 OM-SB 上表面相关 DNA 和蛋白质的测定。还评估了抗菌肽多粘菌素 B 与 OM-SB 的相互作用。这些发现强调了我们的平台能够产生细菌外膜的平面表面的能力,这反过来又可以作为简化抗生素开发的有价值的工具。