Structural Analysis of Multidrug Transport

多药物转运的结构分析

• 批准号: 8197637
• 负责人: MARK E. GIRVIN
• 金额: $ 36.14万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-08-01 至 2014-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8197637
• 关键词: Antibiotics; Bacteria; Binding; Carrier Proteins; Chemical Structure; Complex; Conflict (Psychology); Cytoplasm; Data; Detergents; Disinfectants; Drug Transport; Dyes; Epitopes; Erythromycin; Escherichia coli; Ethidium; Family member; Figs - dietary; Glucosides; Goals; Homo; Hospitals; Ligands; Lipid Bilayers; Lipids; Maps; Membrane; Membrane Proteins; Methods; Micelles; Modeling; Molecular Conformation; Multi-Drug Resistance; Mutagenesis; Paraquat; Peptides; Performance; Pharmaceutical Preparations; Proteins; Protons; Publishing; Pump; Pyronine; Relative (related person); Resistance; Solutions; Staphylococcus aureus; Structural Models; Structure; Sulfadiazine; System; Tetracyclines; Water; Work; antiporter; crosslink; design; inhibitor/antagonist; monomer; multidrug transport; protonation; public health relevance; rhodamine 6G; three dimensional structure

DESCRIPTION (provided by applicant): Bacteria have developed several methods to resist the lethal effects of antibiotics. The broadest spectrum resistance results from the action of Multi-Drug Resistance pumps (MDRs), which extrude a range of compounds of quite diverse chemical structure. The Small Multi-Drug Resistance pumps (SMRs) are 100-110 residue, dimeric proton-drug antiporters that contain the full multidrug transport machinery, stripped to its barest essentials. Hence they are ideal transporters for a comprehensive structural and functional understanding of drug transport and inhibition in a medically important MDR. Unfortunately, the conformational plasticity that enables these proteins to transport such diverse substrates has led to very different structural models and structural interpretations, particularly for the protein in detergent micelles. Thus we propose to determine the lipid bilayer-embedded structures of the conformations making up the functional cycle of the SMR from Staph aureus that provides resistance against common disinfectants, and identify the binding determinants for multiple drugs and inhibitors using solution NMR by: 1) Developing a high performance discoidal peptide-lipid bilayer "nanodiskette" system for NMR studies of membrane proteins, 2) Establishing the relative topology for the Smr monomers (parallel vs inverted), followed by full structure determination of the apo-form of the protein in nanodiskettes, 3) Identifying the binding determinants for transportable drugs and inhibitors, 4) Defining the structural changes in the substrate-bound, and inhibitor-bound Smr transporter, and 5) Characterizing the fold of the most controversial family member, EmrE, in lipid nanodiskettes as a function of protonation state and TPP binding. PUBLIC HEALTH RELEVANCE: The Small Multidrug Resistance transporter protein protects strains of Staph aureus that carry it from common hospital disinfectants. We will determine the three dimensional structure of the protein by itself, and in complex with the compounds that it transports, in order to understand how it functions, and ultimately guide the design of inhibitors of the transporter.

描述（由申请人提供）：细菌已经开发出多种方法来抵抗抗生素的致命作用。最广泛的耐药性源自多重耐药泵 (MDR) 的作用，该泵可挤出一系列化学结构相当不同的化合物。小型多药耐药泵 (SMR) 是 100-110 残留物、二聚质子药物反向转运蛋白，包含完整的多药转运机制，仅保留最基本的要素。因此，它们是全面了解医学上重要的 MDR 中药物转运和抑制的结构和功能的理想转运蛋白。不幸的是，使这些蛋白质能够运输如此多种底物的构象可塑性导致了非常不同的结构模型和结构解释，特别是对于洗涤剂胶束中的蛋白质而言。因此，我们建议确定构成金黄色葡萄球菌 SMR 功能循环的脂质双层嵌入结构，该结构对常见消毒剂具有抗性，并使用溶液 NMR 确定多种药物和抑制剂的结合决定因素：1) 开发用于膜蛋白 NMR 研究的高性能盘状肽-脂质双层“纳米磁盘”系统，2) 建立 Smr 单体的相对拓扑（平行与平行）倒置），然后对纳米软盘中蛋白质的 apo 形式进行完整结构测定，3）识别可转运药物和抑制剂的结合决定因素，4）定义底物结合和抑制剂结合的 Smr 转运蛋白的结构变化， 5) 将脂质纳米盘中最具争议的家族成员 EmrE 的折叠特征描述为质子化状态和 TPP 结合的函数。 公共健康相关性：小型多药耐药转运蛋白可保护携带该蛋白的金黄色葡萄球菌菌株免受常见医院消毒剂的侵害。我们将确定蛋白质本身以及与其转运的化合物复合的三维结构，以了解其功能，并最终指导转运蛋白抑制剂的设计。