The ATP-binding cassette transporter P-glycoprotein (P-gp) is a multidrug efflux pump that is overexpressed in a variety of cancers and associated with the drug-resistance phenomenon. P-gp structures were previously determined in detergent and in nanodiscs, in which different transmembrane helix conformations were found, “straight” and “kinked,” respectively, indicating a possible role of the lipid environment on the P-gp structural ensemble. Here, we investigate the dynamic conformational ensembles and protein-lipid interactions of two human P-gp inward-open conformers, straight and kinked, employing all-atom molecular dynamics (MD) simulations in asymmetric multicomponent lipid bilayers that mimic the highly specialized hepatocyte membrane in which P-gp is expressed. The two conformers are found to differ in terms of the accessibility of the substrate cavity. The MD simulations show how cholesterol and different lipid species wedge, snorkel, and partially enter into the cavity of the straight P-gp conformer solved in detergent. However, access to the cavity of the kinked P-gp conformer solved in nanodiscs is restricted. Furthermore, the volume and dynamic fluctuations of the substrate cavity largely differ between the two P-gp conformers and are modulated by the presence (or absence) of cholesterol in the membrane and/or of ATP. From the mechanistic perspective, the findings indicate that the straight conformer likely precedes the kinked conformer in the functional working cycle of P-gp, with the latter conformation representing a post substrate-bound state. The inaccessibility of the main transmembrane cavity in the kinked conformer might be crucial in preventing substrate disengagement and transport withdrawal. Remarkably, in our unbiased MD simulations, one transmembrane helix (TM10) of the straight conformer underwent a spontaneous transition to a kinked conformation, underlining the relevance of both conformations in a native phospholipid environment and revealing structural descriptors defining the transition between the two P-gp conformers.

中文翻译：

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将脂质的作用与 ABC 转运蛋白 P-糖蛋白的构象动力学耦合


ATP 结合盒转运蛋白 P-糖蛋白 (P-gp) 是一种多药外排泵，在多种癌症中过度表达，并与耐药现象相关。先前在去污剂和纳米圆盘中确定了 P-gp 结构，其中发现了不同的跨膜螺旋构象，分别为“直”和“扭结”，表明脂质环境对 P-gp 结构整体的可能作用。在这里，我们研究了两种人类 P-gp 内向开放构象异构体（直的和扭结的）的动态构象整体和蛋白质-脂质相互作用，在模拟高度专业化的肝细胞膜的不对称多组分脂质双层中采用全原子分子动力学（MD）模拟其中表达P-gp。发现这两种构象异构体在基底腔的可接近性方面有所不同。 MD 模拟显示胆固醇和不同的脂质种类如何楔入、潜行并部分进入溶解在洗涤剂中的直 P-gp 构象异构体的空腔中。然而，进入纳米圆盘中解决的扭结 P-gp 构象异构体的空腔受到限制。此外，两种 P-gp 构象异构体之间的底物腔的体积和动态波动有很大差异，并且受到膜中胆固醇和/或 ATP 的存在（或不存在）的调节。从机制的角度来看，研究结果表明，在 P-gp 的功能工作循环中，直构象异构体可能先于扭结构象异构体，后者构象代表底物结合后的状态。扭结构象异构体中主要跨膜腔的不可接近可能对于防止底物脱离和运输撤回至关重要。 值得注意的是，在我们的无偏 MD 模拟中，直构象异构体的一个跨膜螺旋 (TM10) 自发转变为扭结构象，强调了两种构象在天然磷脂环境中的相关性，并揭示了定义两个 P-构象之间转变的结构描述符。 gp 构象异构体。