Paracini Nicoló, Lakey Jeremy H, Clifton Luke A
Institut Laue-Langevin, Large Scale Structures Group, 71 Avenue des Martyrs, Grenoble 38000, France.
Biosciences Institute, Faculty of Medical Sciences, Newcastle University, Newcastle upon Tyne NE2 4HH, U.K.
ACS Omega. 2025 Jan 14;10(3):2616-2627. doi: 10.1021/acsomega.4c07648. eCollection 2025 Jan 28.
The lipid matrix of the outer membrane (OM) of Gram-negative bacteria consists of a highly asymmetric lipid bilayer containing phospholipids on the inner leaflet and lipopolysaccharides (LPS) in the outer layer. The latter ensures that harmful molecules do not permeate the bacterial cell, but polymyxin B (PmB), a last-resort antibiotic, is capable of interfering with the stability of the LPS layer and overcoming the OM barrier. We have previously shown that the efficacy of PmB in disrupting isotopically asymmetric OM models (H-phospholipids and H-LPS) is regulated by the gel-to-fluid phase transition of the LPS layer. Here, we employ fully deuterated OM models (H-phospholipids and H-LPS) to track the temperature-dependent penetration of PmB within the model membrane by using neutron reflectometry. We use a model-independent approach to quantify PmB penetration as a function of both concentration and temperature as well as a model-dependent analysis to localize PmB in the asymmetric bilayer. By leveraging the ability of neutrons to differentiate hydrogen from deuterium in structural biology we find that PmB hijacks LPS molecules and accumulates predominantly in the hydrophobic region of lipid A.
革兰氏阴性菌外膜（OM）的脂质基质由高度不对称的脂质双层组成，内层小叶含有磷脂，外层含有脂多糖（LPS）。后者确保有害分子不会渗透到细菌细胞中，但多粘菌素B（PmB）作为一种最后的抗生素，能够干扰LPS层的稳定性并克服外膜屏障。我们之前已经表明，PmB破坏同位素不对称外膜模型（H-磷脂和H-LPS）的功效受LPS层凝胶-流体相转变的调节。在这里，我们使用完全氘代的外膜模型（H-磷脂和H-LPS），通过中子反射测量来追踪PmB在模型膜内随温度的渗透情况。我们使用一种与模型无关的方法来量化PmB作为浓度和温度函数的渗透情况，以及一种依赖模型的分析来将PmB定位在不对称双层中。通过利用中子在结构生物学中区分氢和氘的能力，我们发现PmB劫持LPS分子并主要积累在脂质A的疏水区域。
