Molecular Basis of Substrate Translocation in the Drug/H+ Antiporter 1 Family

药物/H 逆向转运蛋白 1 家族底物易位的分子基础

• 批准号: 10414517
• 负责人: Min Lu
• 金额: $ 32.76万
• 依托单位: ROSALIND FRANKLIN UNIV OF MEDICINE & SCI
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-06-15 至 2026-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10414517
• 关键词: Alcoholism; Amino Acids; Antimicrobial Resistance; Binding; Biochemical; Biological; Bipolar Disorder; Brain; Cations; Cell membrane; Cells; Chemical Structure; Clinical; Communicable Diseases; Complex; Coupled; Couples; Data; Development; Drug Binding Site; Drug Efflux; Drug Modelings; Escherichia coli; Family; Gilles de la Tourette syndrome; Goals; Human; Huntington Disease; Infection; Integral Membrane Protein; Lead; Ligands; Light; Major Depressive Disorder; Malignant Neoplasms; Mediating; Membrane; Membrane Proteins; Mental Depression; Modeling; Molecular; Movement; Multi-Drug Resistance; Mutagenesis; Mutate; Neuromodulator; Neurotoxins; Neurotransmitters; Parkinson Disease; Pathogenicity; Patients; Pharmaceutical Preparations; Physiological; Polyamines; Process; Protein Export Pathway; Proteins; Publishing; Reporting; Research; Roentgen Rays; Schizophrenia; Site; Structure; Therapeutic; Transmembrane Transport; Treatment Efficacy; Variant; Work; alcoholism therapy; antimicrobial; antiport; antiporter; autism spectrum disorder; base; clinically relevant; deprotonation; effective therapy; experience; inhibitor; insight; membrane model; microorganism; monoamine; multi drug transporter; mutant; nervous system disorder; neuropsychiatry; neurotransmission; novel strategies; novel therapeutic intervention; overexpression; pathogenic bacteria; protonation; solute; structural biology; targeted treatment; therapeutic target; unpublished works; vesicle transport

Molecular Basis of Substrate Translocation in the Drug/H+ Antiporter 1 Family Summary Integral membrane proteins known as multidrug transporters extrude therapeutic drugs of diverse chemical structures across cell membranes, impeding the treatment of human cancers, infectious diseases, and neurological disorders. Currently we lack a deep and mechanistic understanding of how these proteins export drugs or how they can be thwarted. We will study the structure and mechanism of a model multidrug transporter, MdfA from Escherichia coli, which couples the influx of H+ to the efflux of various antimicrobials and belongs to the ubiquitous Drug/H+ Antiporter 1 (DHA1) family. MdfA orthologues are present in many pathogenic microorganisms, and the overexpression of E. coli MdfA can lead to antimicrobial resistance in clinical patients. Thus, MdfA represents an important target for therapeutic exploitation to overcome multidrug resistance. Furthermore, the SLC18 antiporters, which are the human counterparts of MdfA in the DHA1 family, conduct the H+-dependent vesicular transport of monoamine neurotransmitters, polyamine neuromodulators, and neurotoxins. The human SLC18 antiporters are essential for brain function and promising therapeutic targets for battling alcoholism, autism spectrum disorders, bipolar disorder, Huntington disease, major depressive disorder, Parkinson’s disease, schizophrenia, and Tourette syndrome. Our long-term objective is to understand how the DHA1 multidrug transporters and human SLC18 antiporters translocate their substrates and how their function can be modulated for potential therapeutic benefit. Notably, prior biochemical studies have suggested that MdfA translocates certain substrates via a non-canonical mechanism. Drawing upon these data and our experience in membrane protein structural biology, we will accomplish two aims: (1) to elucidate the molecular basis for simultaneous translocation of two mono-cationic substrates by a DHA1; (2) to reveal the structural mechanism for non-canonical, DHA1- mediated extrusion of di-cationic therapeutics. By combining crystallographic and biochemical studies, we will acquire new insights into how a DHA1 translocates two substrates concurrently, how a DHA1 inhibitor differs from the substrate, and how a DHA1 exports a therapeutic drug in two consecutive and yet different deprotonation/protonation cycles. The new conceptual framework and DHA1 structures obtained from this study will serve as a stepping-stone toward devising novel strategies to evade or inhibit the clinically relevant multidrug transporters, which may rescue therapeutic efficacy against multidrug-resistant cells and halt the spread of untreatable infections. Furthermore, our work will offer a springboard for the mechanistic studies of human SLC18 antiporters, which will shed new light on how they utilize the electrochemical H+ gradient to translocate monoamine neurotransmitters, neurotoxins, or polyamine neuromodulators, across vesicular membranes.

药物/H+逆向转运蛋白1家族底物转位的分子基础 摘要 被称为多药物转运体的整膜蛋白挤出不同种类的治疗药物 穿过细胞膜的化学结构，阻碍人类癌症的治疗，传染性 疾病和神经紊乱。目前，我们缺乏深入和机械性的理解 这些蛋白质输出药物或它们如何被挫败。我们将研究其结构和机制。 一种多药物转运体的模型，来自大肠杆菌的MDFA，它将H+的内流耦合到 是多种抗菌药的外排产物，属于无处不在的药物/H+逆向转运蛋白1(DHA1)家族。 MDFA同源异构体存在于许多致病微生物中，而大肠杆菌的过度表达 MDFA可导致临床患者产生耐药性。因此，千年发展目标代表着一个重要的 治疗利用的目标，以克服多药耐药。此外，SLC18 反转运蛋白是DHA1家族中MDFA的人类对应物，它进行H+依赖的 单胺类神经递质、多胺类神经调节剂和神经毒素的囊泡运输。 人类SLC18逆向转运蛋白对大脑功能是必不可少的，并有望成为治疗癌症的靶点 与酒精中毒、自闭症谱系障碍、双相情感障碍、亨廷顿病、重度抑郁症作斗争 精神障碍、帕金森氏症、精神分裂症和多发性抽动症。我们的长期目标是 了解DHA1多药转运体和人类SLC18反转运体是如何将其 底物以及如何调节它们的功能以获得潜在的治疗益处。值得注意的是，之前 生化研究表明，MDFA通过非规范的 机制。根据这些数据和我们在膜蛋白结构生物学方面的经验，我们 将实现两个目标：(1)阐明两个基因同时易位的分子基础 (2)揭示非正则DHA1-的结构机理。 双阳离子治疗剂的介导性挤出。通过结合结晶学和生物化学研究， 我们将对DHA1如何同时转移两种底物、DHA1如何 抑制剂与底物不同，以及DHA1如何在连续的两个和 然而，不同的去质子化/质子化周期。新的概念框架和DHA1结构 从这项研究中获得的将作为设计新策略的垫脚石，以逃避或 抑制临床相关的多药转运蛋白，可能挽救治疗效果 多药耐药细胞和阻止无法治愈的感染的传播。此外，我们的工作将提供 为人类SLC18逆向转运蛋白的机制研究提供了一个跳板，这将为 他们如何利用电化学H+梯度来转移单胺类神经递质， 神经毒素，或多胺神经调节剂，穿过囊泡膜。