Structure/function analysis of understudied pH-sensitive Kir4.2 channels in vitro with conformation specific nanobodies

使用构象特异性纳米体对正在研究的 pH 敏感 Kir4.2 通道进行体外结构/功能分析

基本信息

  • 批准号:
    10217510
  • 负责人:
  • 金额:
    $ 15.75万
  • 依托单位:
  • 依托单位国家:
    美国
  • 项目类别:
  • 财政年份:
    2021
  • 资助国家:
    美国
  • 起止时间:
    2021-04-15 至 2022-08-14
  • 项目状态:
    已结题

项目摘要

Project Summary Inward rectifying potassium (Kir) channels regulate excitability in many tissues, and multiple diseases result from mutations of Kir channel genes. The long-term goal of this project is to understand the molecular details of understudied pH-sensitive Kir4 sub-family channels. Previously, I defined molecular mechanisms of bulk anionic lipid regulation of Kir2 channels. These previous studies were made possible by my establishing a new expression system and purification protocols for eukaryotic Kir2 channels, as well as uniquely combining the multiple methods of (1) computational docking simulations, (2) functional assays of purified proteins reconstituted in synthetic liposomes, and (3) high resolution structure determination. These studies provide a template for gaining new understanding of understudied pH-sensitive Kir4 channels at the molecular level. We will establish an efficient expression system and a purification protocol for large scale purifications of Kir4.2 homo- and Kir4.2/Kir5.1 hetero-tetramer proteins that are understudied as well as develop nanobodies specific to functional states that will not only facilitate structure determinations in certain functional states but also have therapeutic potentials. Once established, we will utilize the same `triple combination' of approaches to determine molecular details of these understudied channel regulation and function.
项目总结

项目成果

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