Department of Pharmacology, Case Western Reserve University School of Medicine, Cleveland, Ohio, USA.
Department of Chemistry and Biochemistry, University of Oklahoma, Norman, Oklahoma, USA.
mBio. 2021 Feb 23;12(1):e03690-20. doi: 10.1128/mBio.03690-20.
is a Gram-negative pathogen that has emerged as one of the most highly antibiotic-resistant bacteria worldwide. Multidrug efflux within these highly drug-resistant strains and other opportunistic pathogens is a major cause of failure of drug-based treatments of infectious diseases. The best-characterized multidrug efflux system in is the prevalent rug fflux B (AdeB) pump, which is a member of the resistance-nodulation-cell division (RND) superfamily. Here, we report six structures of the trimeric AdeB multidrug efflux pump in the presence of ethidium bromide using single-particle cryoelectron microscopy (cryo-EM). These structures allow us to directly observe various novel conformational states of the AdeB trimer, including the transmembrane region of trimeric AdeB can be associated with form a trimer assembly or dissociated into "dimer plus monomer" and "monomer plus monomer plus monomer" configurations. We also discover that a single AdeB protomer can simultaneously anchor a number of ethidium ligands and that different AdeB protomers can bind ethidium molecules simultaneously. Combined with molecular dynamics (MD) simulations, we reveal a drug transport mechanism that involves multiple multidrug-binding sites and various transient states of the AdeB membrane protein. Our data suggest that each AdeB protomer within the trimer binds and exports drugs independently. has emerged as one of the most highly antibiotic-resistant Gram-negative pathogens. The prevalent AdeB multidrug efflux pump mediates resistance to a broad spectrum of clinically relevant antimicrobial agents. Here, we report six cryo-EM structures of the trimeric AdeB pump in the presence of ethidium bromide. We discover that a single AdeB protomer can simultaneously anchor a number of ligands, and different AdeB protomers can bind ethidium molecules simultaneously. The results indicate that each AdeB protomer within the trimer recognizes and extrudes drugs independently.
是一种革兰氏阴性病原体，已成为全球抗生素耐药性最高的细菌之一。这些高度耐药菌株和其他机会性病原体中的多药外排是导致基于药物的传染病治疗失败的主要原因。 中最具特征的多药外排系统是普遍存在的 rug fflux B（AdeB）泵，它是抗性 - 结节 - 分裂（RND）超家族的成员。在这里，我们使用单颗粒冷冻电子显微镜（cryo-EM）报告了溴化乙锭存在下三聚体 AdeB 多药外排泵的六个结构。这些结构使我们能够直接观察 AdeB 三聚体的各种新颖构象状态，包括三聚体 AdeB 的跨膜区域可以与形成三聚体组装体相关联或解离成“二聚体加单体”和“单体加单体加单体”构型。我们还发现单个 AdeB 原聚体可以同时锚定多个乙锭配体，并且不同的 AdeB 原聚体可以同时结合乙锭分子。结合分子动力学（MD）模拟，我们揭示了一种涉及多个多药结合位点和 AdeB 膜蛋白各种瞬态的药物转运机制。我们的数据表明，三聚体中的每个 AdeB 原聚体都可以独立地结合和输出药物。 已成为最具抗生素耐药性的革兰氏阴性病原体之一。普遍存在的 AdeB 多药外排泵介导对广泛的临床相关抗菌剂的耐药性。在这里，我们报告了溴化乙锭存在下三聚体 AdeB 泵的六个 cryo-EM 结构。我们发现单个 AdeB 原聚体可以同时锚定多个配体，并且不同的 AdeB 原聚体可以同时结合乙锭分子。结果表明，三聚体中的每个 AdeB 原聚体都独立地识别和排出药物。
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