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

基本信息

批准号：
10644018
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
项目类别：
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/10644018
关键词：
Alcoholism Amino Acids Antimicrobial Resistance Binding Biochemical Biological Bipolar Disorder Brain Cell membrane Cells Chemical Structure Clinical Communicable Diseases Complex Coupled Couples Crystallography 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 Ligand Binding Ligands 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 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overexpression pathogenic bacteria protonation solute structural biology targeted treatment therapeutic target unpublished works vesicle transport

项目摘要

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+反向转运蛋白 1 家族底物易位的分子基础概括称为多药转运蛋白的整合膜蛋白可挤出多种治疗药物跨细胞膜的化学结构，阻碍人类癌症、传染病的治疗目前我们对如何发生缺乏深入和机械的理解。我们将研究这些蛋白质输出药物的结构和机制。来自大肠杆菌的多药转运蛋白模型 MdfA，它将 H+ 的流入与多种抗菌药物的外排，属于普遍存在的药物/H+逆向转运蛋白 1 (DHA1) 家族。 MdfA 直系同源物存在于许多病原微生物中，并且大肠杆菌的过度表达 MdfA可导致临床患者产生抗菌药物耐药性，因此，MdfA代表了一个重要的因素。此外，SLC18 是克服多药耐药性的治疗开发目标。反向转运蛋白是 DHA1 家族中 MdfA 的人类盟友，负责 H+ 依赖性单胺神经递质、多胺神经调节剂和神经毒素的囊泡运输。人类 SLC18 反向转运蛋白对于大脑功能至关重要，也是有希望的治疗靶点酗酒、自闭症谱系障碍、双相情感障碍、亨廷顿病、重度抑郁症我们的长期目标是治疗帕金森病、精神分裂症和抽动秽语综合症。了解 DHA1 多药转运蛋白和人类 SLC18 反向转运蛋白如何易位值得注意的是，优先考虑底物以及如何调节它们的功能以获得潜在的治疗益处。生化研究表明，MdfA 通过非规范的方式易位某些底物利用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）阐明两个同时易位的分子基础 DHA1 的单阳离子底物；(2) 揭示非经典 DHA1- 的结构机制通过结合晶体学和生化研究，介导的双阳离子疗法的挤出。我们将获得关于 DHA1 如何同时易位两种底物的新见解，DHA1 如何抑制剂与底物的不同，以及 DHA1 如何在连续和连续的两个过程中输出治疗药物但不同的去质子化/质子化循环。新的概念框架和 DHA1 结构。从这项研究中获得的信息将作为制定新策略来逃避或逃避的踏脚石抑制临床相关的多药转运蛋白，这可能会挽救针对此外，我们的工作还将提供多重耐药细胞并阻止无法治疗的感染的传播。人类 SLC18 反向转运蛋白机制研究的跳板，这将为研究提供新的线索他们如何利用电化学 H+ 梯度来转移单胺神经递质，神经毒素或多胺神经调节剂穿过囊泡膜。