INOSITOL 1,4,5 TRIPHOSPHATE RECEPTOR (IP3R)

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 细胞内钙释放通道是将细胞内钙离子从内质/肌浆网(ER/SR)动员到细胞质中所必需的通道。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，都依赖于细胞质钙离子浓度的瞬时变化[1]。这项建议集中于肌醇1，4，5-三磷酸受体(IP3Rs)，几乎所有细胞的ER/SR膜上都存在同源四聚体钙释放通道。IP3是IP3R的主要配体，与钙离子协同作用，促进IP3通道开放。 小脑浦肯野细胞内质网的主要异构体--1型IP3R(IP3R1)是本研究的重点。IP3R1在小脑ER中形成~1.3个丙二醛的同质异构体[2]。IP3R的结构研究一直受到其大小和与天然构象中的膜脂相互作用的阻碍。由于这种双重复杂性，X射线结晶学和核磁共振波谱不太适合研究大量的完整膜蛋白。在这个项目中，我们的目标是在亚纳米分辨率下确定IP3R1通道的3D结构，并描述其门控的结构决定因素。这种结构信息最终是开发药物策略所必需的，以便在疾病状态下干扰通道蛋白。在我们的结构研究中，我们利用了洗涤剂溶解的纯化通道蛋白的单粒子电子冷冻显微镜和计算机重建技术。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. 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 [1]. 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 [2]. 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.