KCNH channel regulation by intracellular ligands

细胞内配体对 KCNH 通道的调节

• 批准号: 10372601
• 负责人: Tinatin I Brelidze
• 金额: $ 5.63万
• 依托单位: GEORGETOWN UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-05-01 至 2023-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10372601
• 关键词: Arrhythmia; Binding; C-terminal; Cardiac; Cyclic Nucleotides; Development; Electrodes; Electrophysiology (science); Epilepsy; Family; Goals; Ion Channel; Knowledge; Ligands; Malignant Neoplasms; Methods; Molecular; Molecular Mechanisms of Action; Mutagenesis; N-terminal; Pharmacologic Substance; Pharmacology; Physiological; Property; Regulation; Structural Models; Surface Plasmon Resonance; X-Ray Crystallography; base; neuronal excitability; novel; novel strategies; patch clamp; screening; small molecule; small molecule libraries; tool; tumor progression; voltage clamp

Ion channels in the KCNH family perform diverse and important physiological functions, including the regulation of cancer progression, neuronal excitability and cardiac contraction. KCNH channels contain a Per-Arnt-Sim (PAS) domain in their N-terminal and cyclic nucleotide-binding homology (CNBH) domain in their C-terminal regions. These intracellular domains are the key functional domains that are largely responsible for the distinct properties of KCNH channels that determine their physiological applications. The goal of this proposal is to identify small molecule ligands that directly bind to the PAS and CNBH domains and uncover the mechanisms of their action on KCNH channels. We will identify and functionally characterize KCNH channel ligands using a novel strategy based on the combination of surface plasmon resonance (SPR) and electrophysiology as the principal methods. We will first identify small molecule ligands by screening libraries of small molecules against the PAS and CNBH domains of KCNH channels with the high-throughput SPR method. We will then determine the functional effects of the identified small molecule ligands with medium-throughput two- electrode voltage-clamp method. Finally to uncover molecular mechanisms of action of the identified ligands we will use a combination of patch-clamp current recordings, mutagenesis, X-ray crystallography, SPR and structural modeling. These studies will advance our knowledge of KCNH channel regulation by intracellular ligands and PAS and CNBH domains, provide pharmacological tools to study physiological contributions of KCNH channels and will greatly facilitate the development of novel pharmaceutical agents for treatment of cardiac arrhythmias, cancer and epilepsy.

KCNH家族中的离子通道执行多种多样，重要的生理功能， 包括调节癌症进展，神经元兴奋性和心脏 收缩。 KCNH通道在其N末端包含一个均SIM（PAS）域 在其C末端区域中的环状核苷酸结合同源（CNBH）结构域。 这些细胞内域是主要负责的关键功能域 对于确定其生理的KCNH通道的独特特性 申请。该建议的目的是识别直接的小分子配体 与PA和CNBH结构域结合，并揭示其作用机制 KCNH频道。我们将识别并在功能上表征KCNH通道配体 使用基于表面等离子体共振（SPR）组合的新型策略 和电生理作为主要方法。我们将首先识别小分子 配体通过筛选小分子对PA和CNBH结构域的库 具有高通量SPR方法的KCNH通道。然后，我们将确定 鉴定出的小分子配体的功能效应，具有中等通量的两种 电极电压钳法。最终发现了分子机制的作用 所识别的配体我们将使用斑块夹电流记录的组合， 诱变，X射线晶体学，SPR和结构建模。这些研究会 通过细胞内配体和PAS提高我们对KCNH通道调节的了解 和CNBH领域，提供了研究生理贡献的药理工具 KCNH频道的范围，将极大地促进新型药物的发展 治疗心律不齐，癌症和癫痫的药物。