DataSheet1_Bacterial extracellular vesicles: towards realistic models for bacterial membranes in molecular interaction studies by surface plasmon resonance.PDF

One way to mitigate the ongoing antimicrobial resistance crisis is to discover and develop new classes of antibiotics. As all antibiotics at some point need to either cross or just interact with the bacterial membrane, there is a need for representative models of bacterial membranes and efficient methods to characterize the interactions with novel molecules -both to generate new knowledge and to screen compound libraries. Since the bacterial cell envelope is a complex assembly of lipids, lipopolysaccharides, membrane proteins and other components, constructing relevant synthetic liposome-based models of the membrane is both difficult and expensive. We here propose to let the bacteria do the hard work for us. Bacterial extracellular vesicles (bEVs) are naturally secreted by Gram-negative and Gram-positive bacteria, playing a role in communication between bacteria, as virulence factors, molecular transport or being a part of the antimicrobial resistance mechanism. bEVs consist of the bacterial outer membrane and thus inherit many components and properties of the native outer cell envelope. In this work, we have isolated and characterized bEVs from one Escherichia coli mutant and three clinical strains of the ESKAPE pathogens Klebsiella pneumoniae, Acinetobacter baumannii, and Pseudomonas aeruginosa. The bEVs were shown to be representative models for the bacterial membrane in terms of lipid composition with speciesstrain specific variations. The bEVs were further used to probe the interactions between bEV and antimicrobial peptides (AMPs) as model compounds by Surface Plasmon Resonance (SPR) and provide proof-of-principle that bEVs can be used as an easily accessible and highly realistic model for the bacterial surface in interaction studies. This further enables direct monitoring of the effect induced by antibiotics, or the response to host-pathogen interactions.

缓解持续进行的抗菌素耐药性危机的一种途径，在于发现和开发新型抗菌素。鉴于所有抗菌素在某一阶段均需穿越或与细菌细胞膜发生相互作用，构建具有代表性的细菌细胞膜模型和高效表征新分子与细胞膜相互作用的手段显得尤为必要——这不仅有助于产生新的知识，而且可以筛选化合物库。由于细菌细胞外膜是由脂质、脂多糖、膜蛋白及其他成分构成的复杂组装体，因此构建相关的合成脂质体制膜模型既困难又昂贵。在此，我们提出让细菌为我们完成这项艰巨的工作。细菌胞外囊泡（bEVs）由革兰氏阴性和革兰氏阳性细菌自然分泌，其在细菌间的通讯、作为致病因素、分子运输或构成抗菌素耐药机制的一部分中发挥重要作用。bEVs由细菌外膜构成，因而继承了原生外细胞膜中的许多成分和特性。在本研究中，我们已从一株大肠杆菌突变体和三种临床分离株（克雷伯菌肺炎、鲍曼不动杆菌和铜绿假单胞菌）中分离和表征了bEVs。结果显示，bEVs在脂质组成方面是细菌细胞膜的代表性模型，存在物种和菌株特异性变化。进一步地，我们利用表面等离子体共振（SPR）技术，以bEVs作为模型化合物探究了bEVs与抗菌肽（AMPs）之间的相互作用，并提供了原理解证，即bEVs可以作为易于获取且高度逼真的细菌表面模型用于相互作用研究。这进一步使得直接监测抗菌素诱导的效果或宿主-病原体相互作用反应成为可能。