STRUCTURE/FUNCTION STUDIES OF GAP JUNCTIONS

间隙连接的结构/功能研究

• 批准号: 6266215
• 负责人: Thaddeus Andrew Bargiello
• 金额: $ 37.78万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-02-01 至 2004-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6266215
• 关键词: Xenopus oocyte; alternatives to animals in research; aminoacid; biophysics; chimeric proteins; computer simulation; conformation; electric field; electrophysiology; gap junctions; gene mutation; hereditary motor and sensory neuropathy; membrane channels; membrane proteins; nuclear magnetic resonance spectroscopy; protein structure function; site directed mutagenesis; transfection; voltage gated channel

Gap junctions formed by Cx26 and Cx32 differ in molecular permeability and in sensitivity to transjunctional voltage. These differences will be exploited to define the molecular mechanisms that underlie voltage dependence and ionic fluxes of intercellular and membrane channels formed by proteins of the connexin gene family. In the long term, integration of data from biophysical, molecular genetic, NMR and computer modeling studies will lead to understanding of how conformational changes in this membrane protein and others dynamically relate to their function and how fixed charges modulate ion flux. In the proposed studies the pore-lining sequence of the Cx32 channel will be further defined and the atomic structure of the amino terminus of wild type and mutated Cx32 will be solved by high resolution NMR. The position of the voltage sensor and its relation to the transjunctional electric field will be explored. Studies are proposed to test the hypothesis that the conformational flexibility of a proline kink motif underlies the conformational changes required for voltage dependent gating. Studies are also proposed to distinguish between concerted and individual subunit gating models. Data from molecular and electrophysiological studies of ionic flux will be examined using the electrodiffusive model of Chen and Eisenberg to further define the roles of charged amino acids in determining permeation and selectivity of gap junction channels. X-linked Charcot-Marie-Tooth disease (CMTX) is caused by mutations of human Cx32, and nonsyndromic deafness by mutations of Cx26. Biophysical and molecular studies have shown that changes in permeability of Cx32 are likely to underlie the etiology of CMTX. The proposed studies will advance the basic scientific knowledge required to define the molecular basis of this and other connexin-related diseases, and the biological role of gap junctions.

由Cx26和Cx32形成的间隙连接在分子水平上不同， 渗透性和对跨接电压的敏感性。这些差异 将被用来定义电压的分子机制 依赖性和离子通量的细胞间和膜通道形成的 连接蛋白基因家族的蛋白质。从长远来看，数据集成 从生物物理学、分子遗传学、核磁共振和计算机建模研究， 从而了解这种膜蛋白的构象变化 而其他的则与它们的功能以及固定电荷如何调节 离子通量在所提出的研究中，Cx32通道的孔衬序列 将被进一步定义，野生型的氨基末端的原子结构 型和突变的Cx32将通过高分辨率NMR来解决。的位置 电压传感器及其与过渡电场的关系将 被探索。研究提出了测试的假设， 脯氨酸扭结基序的构象灵活性是构象的基础。 电压相关门控所需的变化。还建议进行研究， 区分协同和个体亚基门控模型。数据从 离子通量的分子和电生理学研究将使用 Chen和Eisenberg的电扩散模型，以进一步定义 带电荷氨基酸测定间隙连接的渗透性和选择性 渠道X连锁腓骨肌萎缩症（CMTX）是由以下基因突变引起的： 人Cx32和Cx26突变引起的非综合征性耳聋。生物物理和 分子研究表明，Cx32渗透性的变化可能 CMTX的病因。拟议的研究将推进基本的 需要科学知识来定义这种和其他的分子基础 连接蛋白相关疾病，以及缝隙连接的生物学作用。