L-type Ca2+ Channel Regulation by Calmodulin and CaBP1

钙调蛋白和 CaBP1 对 L 型 Ca2 通道的调节

基本信息

批准号：
10618394
负责人：
JAMES B AMES
金额：
$ 31.4万
依托单位：
UNIVERSITY OF CALIFORNIA AT DAVIS
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-06-01 至 2025-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10618394
关键词：
Actinin Alzheimer&apos s Disease Binding Binding Sites Biological Assay Brain C-terminal CTF1 gene Calcium Calcium Binding Calcium Channel Calcium Signaling Calcium ion Calcium-Binding Proteins Calmodulin Cells Complex Computer Analysis Cryoelectron Microscopy Dissociation EF Hand Motifs Electrophysiology (science)Epilepsy Feedback Fluorescence Gene Expression Goals Heart Heterogeneity Ion Channel Learning Link Lobe Mediating Memory Migraine Modeling Molecular Conformation Molecular Structure Mutation Mutation Analysis Neurons Nuclear Magnetic Resonance Physiological Postsynaptic Membrane Proteins Regulation Research Retina Role Sampling Signal Transduction Site-Directed Mutagenesis Structure Techniques Testing Timothy syndrome Work X-Ray Crystallography chronic pain in vivo insight microcalorimetry mutant nervous system disorder neuronal excitability neurotransmission postsynaptic neurons prevent response sensor transmission process voltage

项目摘要

The overall objectives are to apply nuclear magnetic resonance (NMR) techniques in concert with experimental functional approaches to elucidate the molecular structure and regulatory mechanisms of neuronal L-type voltage-gated Ca2+ channel (CaV1.2) and its atomic-level structural and functional association with calcium binding protein-1 (CaBP1), -actinin1 (ACTN1) and calmodulin (CaM). During the next five years, we will use NMR, fluorescence, microcalorimetry, cryoEM, x-ray crystallography, and computational analysis to delineate the structure and dynamics of CaM and CaBP1 each bound to CaV1.2. Site-directed mutagenesis and electrophysiology functional analysis will be integrated with atomic– level structural information to understand how CaM, ACTN1 and CaBP1 each reciprocally control the Ca2+- dependent channel activity of CaV1.2 in neuronal functions. By pursuing these studies, we hope to gain an atomic-level understanding of how CaM, ACTN1 and CaBP1 each regulate CaV1.2 involved in controlling neuronal excitability and gene expression. In particular, we want to understand how protein target binding sites work in concert with calcium-binding sites to confer Ca2+-dependent channel activity of CaV1.2 at the postsynaptic membrane. This structural information will probe how L-type channel regulation may be connected to neurological disorders, including epilepsy, Alzheimer’s disease, and Timothy Syndrome. The Specific Aims are three-fold: (1) Determine the structural basis of CaV1.2 channel activation promoted by CaM and ACTN1; (2) Elucidate the structure and functional role of a CaM intermediate with 2 Ca2+ bound and determine its role in regulating Ca2+-dependent inactivation (CDI); (3) Determine the structural basis of how CaBP1 suppresses Ca2+-dependent inactivation (CDI) of CaV1.2.

总体目标是将核磁共振（NMR）技术应用于与实验功能方法的音乐会，以阐明分子结构和调节神经元L型电压门控Ca2+通道（CAV1.2）及其原子级结构和与钙结合蛋白-1（CABP1），-actinin1（ACTN1）和钙调蛋白（CAM）的功能关联。在接下来的五年中，我们将使用NMR，荧光，微钙化，冷冻，X射线晶体学，，和计算分析以描述CAM和CABP1的结构和动力学 Cav1.2。位置定向的诱变和电生理功能分析将与原子 - 级别的结构信息以了解CAM，ACTN1和CABP1如何相互控制Ca2+ - CAV1.2在神经元功能中的依赖通道活性。通过追求这些研究，我们希望获得原子级对CAM，ACTN1和CABP1的了解如何调节CAV1.2控制神经元令人兴奋和基因表达。特别是，我们想了解蛋白质的目标绑定位点与钙结合位点共同与会议Ca2+依赖性通道活动的CAV1.2合作。在突触后膜上。这些结构信息将探测L型通道调节的调节与神经系统疾病有关，包括癫痫，阿尔茨海默氏病和蒂莫西综合症。具体目的是三倍：（1）确定促进CAV1.2通道激活的结构基础由CAM和ACTN1; （2）阐明了与2个Ca2+的CAM中间体的结构和功能作用绑定并确定其在确定Ca2+依赖性失活（CDI）中的作用；（3）确定结构 CABP1如何抑制Cav1.2的Ca2+依赖性失活（CDI）的基础。