Ilangumaran Ponmalar Ilanila, Sarangi Nirod K, Basu Jaydeep K, Ayappa K Ganapathy
Center for BioSystems Science and Engineering, Indian Institute of Science, Bangalore, India.
School of Chemical Science, Dublin City University, Dublin, Ireland.
Front Mol Biosci. 2021 Sep 9;8:737561. doi: 10.3389/fmolb.2021.737561. eCollection 2021.
Pore forming proteins are a broad class of pathogenic proteins secreted by organisms as virulence factors due to their ability to form pores on the target cell membrane. Bacterial pore forming toxins (PFTs) belong to a subclass of pore forming proteins widely implicated in bacterial infections. Although the action of PFTs on target cells have been widely investigated, the underlying membrane response of lipids during membrane binding and pore formation has received less attention. With the advent of superresolution microscopy as well as the ability to carry out molecular dynamics (MD) simulations of the large protein membrane assemblies, novel microscopic insights on the pore forming mechanism have emerged over the last decade. In this review, we focus primarily on results collated in our laboratory which probe dynamic lipid reorganization induced in the plasma membrane during various stages of pore formation by two archetypal bacterial PFTs, cytolysin A (ClyA), an -toxin and listeriolysin O (LLO), a -toxin. The extent of lipid perturbation is dependent on both the secondary structure of the membrane inserted motifs of pore complex as well as the topological variations of the pore complex. Using confocal and superresolution stimulated emission depletion (STED) fluorescence correlation spectroscopy (FCS) and MD simulations, lipid diffusion, cholesterol reorganization and deviations from Brownian diffusion are correlated with the oligomeric state of the membrane bound protein as well as the underlying membrane composition. Deviations from free diffusion are typically observed at length scales below ∼130 nm to reveal the presence of local dynamical heterogeneities that emerge at the nanoscale-driven in part by preferential protein binding to cholesterol and domains present in the lipid membrane. Interrogating the lipid dynamics at the nanoscale allows us further differentiate between binding and pore formation of - and -PFTs to specific domains in the membrane. The molecular insights gained from the intricate coupling that occurs between proteins and membrane lipids and receptors during pore formation are expected to improve our understanding of the virulent action of PFTs.
成孔蛋白是一类广泛存在的致病蛋白，由于其能够在靶细胞膜上形成孔道，生物体将其作为毒力因子分泌出来。细菌成孔毒素（PFTs）属于成孔蛋白的一个亚类，广泛参与细菌感染。尽管PFTs对靶细胞的作用已得到广泛研究，但在膜结合和孔形成过程中脂质的潜在膜反应却较少受到关注。随着超分辨率显微镜的出现以及对大型蛋白质膜组装体进行分子动力学（MD）模拟的能力，在过去十年中出现了关于成孔机制的新颖微观见解。在本综述中，我们主要关注我们实验室整理的结果，这些结果探究了两种典型细菌PFTs，即溶血素A（ClyA）（一种α毒素）和李斯特菌溶血素O（LLO）（一种β毒素）在孔形成的各个阶段诱导质膜中动态脂质重排的情况。脂质扰动的程度既取决于孔复合体插入膜的基序的二级结构，也取决于孔复合体的拓扑变化。使用共聚焦和超分辨率受激发射损耗（STED）荧光相关光谱（FCS）以及MD模拟，脂质扩散、胆固醇重排以及与布朗扩散的偏差与膜结合蛋白的寡聚状态以及潜在的膜组成相关。通常在长度尺度低于约130 nm时观察到与自由扩散的偏差，以揭示局部动态异质性的存在，这种异质性在纳米尺度上出现，部分是由蛋白质与胆固醇和脂质膜中存在的结构域的优先结合驱动的。在纳米尺度上研究脂质动力学使我们能够进一步区分α - 和β - PFTs与膜中特定结构域的结合和孔形成。在孔形成过程中蛋白质与膜脂质和受体之间发生的复杂耦合所获得的分子见解，有望增进我们对PFTs毒力作用的理解。
