PfMDR1: Mechanisms of Transport Modulation by Functional Polymorphisms

ATP-Binding Cassette (ABC) transporters are efflux pumps frequently associated with multidrug resistance in many biological systems, including malaria. Antimalarial drug-resistance involves an ABC transporter, PfMDR1, a homologue of P-glycoprotein in humans. Twenty years of research have shown that several single nucleotide polymorphisms in pfmdr1 modulate in vivo and/or in vitro drug susceptibility. The underlying physiological mechanism of the effect of these mutations remains unclear. Here we develop structural models for PfMDR1 in different predicted conformations, enabling the study of transporter motion. Such analysis of functional polymorphisms allows determination of their potential role in transport and resistance. The bacterial MsbA ABC pump is a PfMDR1 homologue. MsbA crystals in different conformations were used to create PfMDR1 models with Modeller software. Sequences were aligned with ClustalW and analysed by Ali2D revealing a high level of secondary structure conservation. To validate a potential drug binding pocket we performed antimalarial docking simulations. Using aminoquinoline as probe drugs in PfMDR1 mutated parasites we evaluated the physiology underlying the mechanisms of resistance mediated by PfMDR1 polymorphisms. We focused on the analysis of well known functional polymorphisms in PfMDR1 amino acid residues 86, 184, 1034, 1042 and 1246. Our structural analysis suggested the existence of two different biophysical mechanisms of PfMDR1 drug resistance modulation. Polymorphisms in residues 86/184/1246 act by internal allosteric modulation and residues 1034 and 1042 interact directly in a drug pocket. Parasites containing mutated PfMDR1 variants had a significant altered aminoquinoline susceptibility that appears to be dependent on the aminoquinoline lipophobicity characteristics as well as vacuolar efflux by PfCRT. We previously described the in vivo selection of PfMDR1 polymorphisms under antimalarial drug pressure. Now, together with recent PfMDR1 functional reports, we contribute to the understanding of the specific structural role of these polymorphisms in parasite antimalarial drug response.

ATP结合盒（ATP-Binding Cassette, ABC）转运蛋白是一类外排泵，在包括疟疾在内的诸多生物系统中，常与多药耐药性密切相关。抗疟药性涉及一种ABC转运蛋白PfMDR1，它是人类P-糖蛋白（P-glycoprotein）的同源物。二十余年的研究表明，pfmdr1基因中的若干单核苷酸多态性（single nucleotide polymorphisms）会调控体内和/或体外的药物敏感性，但此类突变发挥作用的潜在生理机制至今仍不明确。本研究构建了不同预测构象下的PfMDR1结构模型，为解析该转运蛋白的运动机制提供了可行路径。针对功能多态性的此类分析，可用于确定其在转运过程与耐药性中的潜在作用。细菌MsbA ABC泵是PfMDR1的同源蛋白，研究人员借助Modeller软件，以不同构象下的MsbA晶体结构为模板构建PfMDR1模型。序列通过ClustalW进行比对，并经Ali2D分析，结果显示其二级结构具有高度保守性。为验证潜在的药物结合口袋，我们开展了抗疟药物分子对接模拟（docking simulations）。以氨基喹啉（aminoquinoline）类药物作为探针，在携带PfMDR1突变的疟原虫中评估了由PfMDR1多态性介导的耐药性相关生理机制。本研究聚焦于PfMDR1第86、184、1034、1042及1246位氨基酸残基处的已知功能多态性分析。结构分析结果表明，PfMDR1的耐药性调控存在两种不同的生物物理机制：第86/184/1246位残基的多态性通过内部变构调控（allosteric modulation）发挥作用，而第1034与1042位残基则直接参与药物口袋的相互作用。携带突变型PfMDR1变体的疟原虫，其氨基喹啉类药物敏感性发生显著改变，这种改变似乎与氨基喹啉的疏脂特性以及PfCRT介导的液泡外排过程相关。我们此前曾报道过在抗疟药物压力下体内PfMDR1多态性的筛选情况。如今，结合近期的PfMDR1功能研究报告，本研究有助于进一步阐明这些多态性在疟原虫抗疟药物应答中的特定结构功能。