Structural Dynamics of Multi-drug Resistance ABC Transporters

多药耐药ABC转运蛋白的结构动力学

• 批准号: 7907063
• 负责人: Hassane S Mchaourab
• 金额: $ 16.53万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-31 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7907063
• 关键词: ATP Hydrolysis; ATP-Binding Cassette Transporters; Active Biological Transport; Address; Amino Acid Sequence; Architecture; Binding; Biochemical; Biological Models; Cells; Chemotherapy-Oncologic Procedure; Clinical; Couples; Crystallography; Cystic Fibrosis; Data; Development; Dimensions; Dimerization; Dissociation; Electrons; Energy Metabolism; Energy Transfer; Equilibrium; Escherichia; Fluorescence; Funding; Genome; Goals; Hereditary Disease; Homeostasis; Human; Hydrolysis; Inherited; Label; Ligands; Lipid A; Lipid Bilayers; Lipids; Lipopolysaccharides; Liposomes; Maps; Measurement; Measures; Membrane; Modeling; Molecular Conformation; Morphologic artifacts; Motion; Movement; Multi-Drug Resistance; Mycoses; Nature; Nucleotides; Pathology; Peptide Sequence Determination; Pharmaceutical Preparations; Play; Process; Proteins; Publishing; Pump; Reagent; Relative (related person); Reporter; Request for Proposals; Research; Research Personnel; Resistance; Rest; Role; Scanning; Shapes; Side; Site; Spectrum Analysis; Spin Labels; Stroke; Structure; Techniques; Temperature; Testing; Transmembrane Domain; base; chemotherapy; clinically relevant; cytotoxic; design; dimer; efflux pump; fascinate; flexibility; global environment; luminescence resonance energy transfer; multi drug transporter; novel; periplasm; programs; quantum; solute; therapeutic development; tool

Clinical multidrug resistance in the treatment of bacterial and fungal infections and cancer chemotherapy can result from expression of membrane-embedded efflux pumps that extrude cytotoxic molecules out of the cell. A subclass of these pumps consists of ATP binding cassette (ABC) transporters, ATP-powered traffickers of a wide range of molecules. The long term goal of this research is to define the protein motion that couples energy expenditure to solute translocation by ABC transporters. Roughly 5% of the Escherichia Co//genome encodes for ABC transporters; one of which, MsbA, transduces ATP energy to flip lipid A, the building block of the outer membrane, across the inner membrane. Its critical role in bacterial homeostasis, sequence and functional similarity to human multidrug transporters in conjunction with extensive crystallographic analysis make MsbA a biochemically and biophysically tractable model of clinically relevant ATP-coupled transport. We will use spectroscopic techniques to obtain direct structural and dynamic information on well-defined, key catalytic intermediates of MsbA in lipid bilayers and in the absence of conformational selectivity by crystal lattice forces. Thespecific aims willtest a mechanism of transport that envisions anATP- and substrate- regulated switch whereby transport occurs concomitantly with cycles of dimerization and dissociation of the nucleotide binding domains (NBDs). Spin labels will be systematically introduced into the protein sequence and their mobilities, accessibilites and pairwise proximities analyzed by electron paramagnetic spectroscopy (EPR) to 1) reconstruct the relative movements of the NBDs and 2) map conformational changes that reorient the substrate binding chamber. A novel aspect of this proposal is the use of complementary spectroscopic rulers with a 5-80A distance range to address controversial aspects of the crystal structures and add a dynamic dimension to these static snapshots. The proposed studies will bridge the current divide between structural and mechanistic models of ABC transporters and provide a unique dynamic perspective on a process of fundamental biochemical importance. In addition to controlling the pharmokinetic profile of xenotoxins, human ABC transporters play causative roles in a number of genetic disorders including cystic fibrosis. Understanding their mechanisms will aid in the design of new drugs to overcome resistance to chemotherapy and the development of therapeutic strategies for the inherited pathologies.

临床多药耐药在治疗细菌和真菌感染和癌症化疗中可以 这是由于嵌入膜的外排泵将细胞毒分子挤出细胞外的结果。 这些泵的一个亚类包括三磷酸腺苷结合盒(ABC)转运体，三磷酸腺苷驱动的转运体 各种各样的分子。这项研究的长期目标是定义 ABC转运蛋白对溶质转运的能量消耗。大约5%的大肠杆菌Co//基因组 ABC转运蛋白的编码；其中之一，MSBA，将ATP能量转导到翻转脂质A，这是构建块 在外膜上，穿过内膜。它在细菌动态平衡、序列和 与人类多药转运蛋白的功能相似性结合广泛的结晶学分析 使MSBA成为临床上相关的ATP偶联转运的生物化学和生物物理易于处理的模型。 我们将使用光谱技术来获得关于定义明确的关键字的直接结构和动态信息 MSBA在脂质双层中的催化中间体及其晶体构象选择性的研究 晶格力。这些特定的目标将测试一种运输机制，该机制设想了一种ATP-和底物- 调节的开关，运输伴随着二聚化和解离的循环而发生 核苷酸结合域(NBD)。自旋标记将被系统地引入蛋白质序列 并用电子顺磁共振谱分析了它们的迁移率、可达性和成对邻近度 (EPR)以1)重建NbD的相对运动和2)映射构象变化 调整底物结合室的方向。这一提议的一个新的方面是使用补语 具有5-80A距离范围的尺子光谱，以解决晶体结构中有争议的方面 并为这些静态快照添加动态维度。拟议的研究将弥合目前的鸿沟 ABC运输机的结构模型和机械模型之间的关系，并提供了一个独特的动态视角 在一个具有基本生物化学重要性的过程中。除了控制药物的药动学特征外 异种毒素，人类ABC转运蛋白在许多遗传性疾病中发挥作用，包括囊性 纤维化症。了解它们的机制将有助于设计克服抗药性的新药 针对遗传性病理的化疗和治疗策略的发展。