INOSITOL 1,4,5 TRIPHOSPHATE RECEPTOR (IP3R)

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 细胞内钙释放通道是将钙从细胞内储存库如内质网/肌浆网（ER/SR）动员到细胞质中所必需的。各种各样的细胞过程，包括肌肉收缩、细胞增殖、分泌、受精和细胞分化，依赖于胞质Ca 2+浓度的瞬时变化。该建议集中在肌醇1，4，5-三磷酸受体（IP 3R），几乎所有细胞的ER/SR膜中发生的同源四聚体Ca 2+释放通道。IP 3是IP 3R的主要配体，其与Ca 2+协同促进IP 3通道开放。 1型IP 3R（IP 3R 1）是小脑浦肯野细胞ER的主要亚型，是本项目的重点。IP 3R 1在小脑ER中形成约1.3 MDa的同源四聚体。IP 3Rs的结构研究受到其大小及其与膜脂质在其天然构象中的相互作用的阻碍。由于这种双重复杂性，X射线晶体学和NMR光谱学不太适合研究大量的膜蛋白。在这个项目中，我们的目标是确定亚纳米分辨率的IP 3R 1通道的三维结构，并描绘其门控的结构决定因素。这些结构信息最终需要用于开发在疾病状态下干扰通道蛋白的药理学策略。在我们的结构研究中，我们利用洗涤剂溶解的纯化通道蛋白和计算机重建技术的单粒子电子冷冻显微镜。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Intracellular Ca2+-release channels are essential for mobilizing Ca2+ from intracellular stores such as the endoplasmic/sarcoplasmic reticulum (ER/SR) into the cytoplasm of cells. A wide variety of cellular processes, including muscle contraction, cell proliferation, secretion, fertilization, and cell differentiation, rely on transient changes in cytoplasmic Ca2+ concentration. This proposal focuses in inositol 1,4,5-trisphosphate receptors (IP3Rs), homotetrameric Ca2+-release channels occurring in the ER/SR membranes of virtually all cells. IP3 is the primary IP3R ligand, which synergizes with Ca2+ to promote IP3 channel opening. Type-1 IP3R (IP3R1), the predominant isoform in the ER of cerebellar Purkinje cells, is the focus of this project. IP3R1 forms homotetramers of ~1.3 MDa in the cerebellar ER. Structural studies of IP3Rs have been hampered by both its size and its interaction with membrane lipids in its native conformation. X-ray crystallography and NMR spectroscopy are poorly suited methodologies to study massive integral membrane proteins due to this double complexity. In this project, we aim to determine 3D structure of IP3R1 channel at subnanometer resolution and to delineate structural determinants of its gating. Such structural information is ultimately needed for the development of pharmacological strategies to interfere with the channel protein in disease states. In our structural studies we exploit single particle electron cryomicroscopy of the detergent-solubilized purified channel protein and computer reconstruction techniques.