ION CHANNELS AND OSMOTAXIS IN ESCHERICHIA COLI

大肠杆菌中的离子通道和渗透性

• 批准号: 3238820
• 负责人: JULIUS ADLER
• 金额: $ 18.56万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 1987
• 资助国家: 美国
• 起止时间: 1987-07-15 至 1992-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3238820
• 关键词: Escherichia coli; autoradiography; bacterial genetics; biological signal transduction; cell osmotic pressure; chemotaxis; cilium /flagellum motility; electron microscopy; electrophysiology; gel electrophoresis; gene mutation; genetic manipulation; genetic mapping; immunofluorescence technique; ion transport; laboratory rabbit; membrane activity; membrane channels; membrane model; membrane permeability; membrane potentials; membrane proteins; model design /development; molecular cloning; nucleic acid sequence; osmosis; point mutation; protein sequence; protoplast /spheroplast

Ion channels underlie physiological functions from nerve excitation to blood-cell volume control as well as pathological conditions such as cystic fibrosis and lupus erythromatosis. Osmoregulation is important in animal cells in general and in blood and kidney cells in particular. Both ion channels and osmoregulation evolved early and have been found in the bacterium Escherichia coli. The long-term goal of this projected is to use E. coli to understand the structures and functions of ion channels and the molecular mechanism of osmotaxis. The patch-clamp survey will be continued to uncover the electric currents through different types of ion channels on the inner or on the outer membranes of E. coli. Their unit conductances, ion specificities, gating mechanisms and kinetic properties will be described. To find out the genes that correspond to the ion currents, existing mutants which may code for channels (phoE-, ompA, envZ-, trkB-, trkC-, osmoregulatory mutants, chemotactic mutants) will be examined for their electrophysiological phenotypes. Ion-resistant, ion-sensitive and osmotactic mutants will be isolated and subjected to the same scrutiny. Once the mutations which correspond to the loss of ion currents are identified, they will be grouped and mapped. The genes suspected to code for ion channel structures will be cloned. Structural models will be built based on the amino-acid sequences. Random mutations selected in vivo as well as mutations directed to specific sites in vitro will be induced and their bioelectric consequences analyzed to test the proposed models. To understand osmotaxis, the existing mutants defective in sensory transduction (flaI-, che-, mot-) will be examined for their osmotactic ability. Osmotactic mutants to be isolated will be used to find the genes and gene products used in this taxis. The possibility of repulsion by low osmolarity besides high osmolarity will be explored. The point at which the osmotactic and chemotactic information merge will be identified through mutational analyses and the possibility that taxis and/or motility requires a proper turgor pressure will be examined. The relation between pressure-activated ion channels and osmotaxis will be examined by electrical studies of osmotactic mutants and behavioral studies of the ion-channel mutants.

离子通道是神经生理功能的基础 刺激血细胞容量控制以及病理 例如囊性纤维化和红斑狼疮。 渗透调节在一般动物细胞中是重要的， 尤其是血液和肾细胞 离子通道和 神经调节很早就进化了， 大肠杆菌。 这一计划的长期目标 就是用E.了解离子的结构和功能 通道和趋化的分子机制。 膜片钳技术的研究将继续揭示电生理学中 电流通过不同类型的离子通道的内部或 E.杆菌 它们的单位电导，离子 特异性，门控机制和动力学特性将是 介绍了 找出与离子相对应的基因 电流，可能编码通道的现有突变体（phoE-， ompA、envZ-、trkB-、trkC-、p53调节突变体、趋化性 突变体）将检查其电生理学 表型 离子抗性、离子敏感性和趋化性突变体 都将被隔离并接受同样的审查 一旦 对应于离子电流损失的突变是 一旦确定，将对其进行分组并绘制地图。 怀疑的基因 离子通道结构的编码将被克隆。 结构 将根据氨基酸序列建立模型。 随机 体内选择的突变以及针对 将在体外诱导特定位点， 分析结果以测试所提出的模型。 到 理解趋化性，现有的突变体在感觉上有缺陷， 转导（flaI-，che-，mot-）将被检查其 战术能力。 要分离的渗透调节突变体将是 用来寻找这种出租车中使用的基因和基因产物。 的 除高渗透压外，低渗透压也可能导致排斥 将被探索。 在这一点上， 趋化性信息合并将通过以下方式确定： 突变分析和可能性，出租车和/或运动 将检查是否需要适当的膨压。 的关系 压力激活离子通道和趋化性之间的联系 通过对趋化突变体的电学研究进行了检查， 离子通道突变体的行为研究。