Structure and mechanism of zinc efflux transporters

锌外流转运蛋白的结构和机制

• 批准号: 8450117
• 负责人: Dax Fu
• 金额: $ 30.92万
• 依托单位: JOHNS HOPKINS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-04-01 至 2015-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8450117
• 关键词: Active Sites; Address; Allosteric Regulation; Alzheimer' s Disease; Architecture; Binding; Biochemical; Biological Assay; Budgets; Cations; Chemicals; Cytoplasm; Cytoplasmic Tail; Diabetes Mellitus; Diffusion; Drug Targeting; Environment; Escherichia coli; Event; Family; Family member; Genetic Polymorphism; Genetic Screening; Goals; Homeostasis; Homology Modeling; Human; Ions; Kinetics; Knowledge; Life; Lipids; Membrane; Membrane Transport Proteins; Metals; Methodology; Modeling; Molecular; Molecular Conformation; Movement; Names; Organism; Orthologous Gene; Pathway interactions; Pharmaceutical Preparations; Play; Proteins; Protons; Regulation; Research; Research Project Grants; Resolution; Risk Assessment; Role; Shapes; Solvents; Specificity; Structure; Surface; Techniques; Time; Transmembrane Domain; Transport Reaction; Variant; Work; Zinc; aqueous; base; disorder risk; drug discovery; extracellular; infancy; metal metabolism; millisecond; molecular marker; programs; response; restraint; zinc-binding protein

DESCRIPTION (provided by applicant): Zinc transporters play a central role in metal metabolism and homeostasis in all living organisms. Our long- term goal is to understand the structure and mechanism of zinc transporters and their functional regulation by metal ions, lipids, and drugs. This is the second competitive renewal of a continuing research project focusing on zinc-efflux transporters from the cation diffusion facilitator (CDF) family. In the current budget period, we solved the crystal structure of YiiP, a prototypic CDF ortholog from E. coli. The crystal structure reveals a Y- shaped architecture that is uniquely oriented in the membrane with an active-site situated near the protein-lipid interface. Despite our existing crystallographic snapshots, a structure-based transport model still is lacking, and the relevance of the YiiP crystal structure to its native structure in the membrane is uncertain. Further, YiiP is a two-modular membrane transporter with a cytoplasmic domain for zinc sensing, and a transmembrane domain that transports zinc ions in response to fluctuation of cytoplasmic zinc concentrations. The conformational changes underlying the allosteric regulation of zinc transport are yet to be explored. In the next budget period, we will use a combination of low-resolution biochemical techniques and high-resolution crystallographic analysis to address three questions: (i) how does YiiP move a cytoplasmic zinc ion across the membrane barrier, (ii) how is the active-site reshaped by surrounding lipids, and, (iii) how do conformational changes in YiiP enable a regulated zinc-transport activity tunable to the cytoplasmic zinc concentration? Accordingly, we propose three specific aims: (i) developing a mechanistic model for zinc transport, (ii) evaluating YiiP conformations in the membrane, and, (iii) defining the YiiP structure in action. During the past two budget cycles, we have developed functional assays for zinc transport with millisecond time-resolution, and obtained structural information on a CDF ortholog at atomic resolution. In the next budget period, we will explore the inner workings of CDFs in atomic detail (aim-1), in native membrane environments (aim-2), and in action (aim-3).

描述（由申请人提供）：锌转运蛋白在所有生物体中的金属代谢和稳态中起着核心作用。我们的长期目标是了解金属离子，脂质和药物的锌转运蛋白的结构和机制及其功能调节。这是持续研究项目的第二个竞争续约，该项目重点是阳离子扩散促进剂（CDF）家族的锌 - 效能转运蛋白。在当前的预算期间，我们解决了Yiip的晶体结构，Yiip是大肠杆菌的原型CDF直系同源物。晶体结构揭示了Y形结构，该结构在膜上独特地定向，该膜位于蛋白质 - 脂质界面附近的活动位置。尽管我们现有的晶体学快照，但仍缺乏基于结构的传输模型，Yiip晶体结构与其在膜中的天然结构的相关性尚不确定。此外，YIIP是一种具有细胞质结构域的两种模块化膜转运蛋白，用于锌传感，而跨膜结构域则响应细胞质锌浓度的波动而运输锌离子。锌运输的变构调节的基础构象变化尚未探讨。 In the next budget period, we will use a combination of low-resolution biochemical techniques and high-resolution crystallographic analysis to address three questions: (i) how does YiiP move a cytoplasmic zinc ion across the membrane barrier, (ii) how is the active-site reshaped by surrounding lipids, and, (iii) how do conformational changes in YiiP enable a regulated zinc-transport activity tunable to细胞质锌浓度？因此，我们提出了三个具体的目的：（i）开发锌转运的机械模型，（ii）评估膜中的YIIP构象，以及（iii）定义在作用中的YIIP结构。在过去的两个预算周期中，我们以毫秒的时间分辨率开发了锌运输的功能测定，并在原子分辨率下获得了CDF直系同源物的结构信息。在下一个预算期间，我们将在原子细节（AIM-1），本机膜环境（AIM-2）和行动中（AIM-3）中探索CDF的内部运作。