DETERMINATION OF THE STRUCTURAL BASIS FOR PICK1 REGULATION

确定 PICK1 监管的结构基础

• 批准号: 8363555
• 负责人: ROBERTO DOMINGUEZ
• 金额: $ 1.19万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2012-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8363555
• 关键词: Address; Affect; Banana; Binding; C-terminal; Cellular Membrane; Complex; Cytoplasm; Data; Funding; Grant; Learning; Length; Location; Membrane; Memory; N-terminal; National Center for Research Resources; Neurons; Peptides; Phosphotransferases; Positioning Attribute; Principal Investigator; Process; Propionic Acids; Proteins; Radial; Regulation; Relative (related person); Research; Research Infrastructure; Resources; Shapes; Source; Tail; Tertiary Protein Structure; United States National Institutes of Health; amphiphysin; base; cost; env Gene Products; protein structure; receptor; trafficking

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. Protein interacting with C-kinase 1 (PICK1) is required for ¿-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor trafficking in neurons, a crucial process involved in learning and memory. PICK1 contains an N-terminal PSD-95/Discs-large/ZO-1 (PDZ) domain followed by a central membrane-binding Bin/amphiphysin/Rvs (BAR) domain and a C-terminal acidic domain. PICK1 is regulated by Ca2+. In the absence of Ca2+, PICK1 is localized to the cytoplasm, suggesting that PICK1 is normally auto-inhibited. Upon Ca2+ stimulation PICK1 binds to the AMPA receptor tail through its PDZ domain and to the adjacent membrane via its BAR domain. Thus, a major conformational change is expected to take place between the Ca2+-bound and Ca2+-free states. We propose to use small-angle x-ray scattering (SAXS) to address how Ca2+ binding affects the structure of PICK1. Specifically, we plan to determine the overall envelope of PICK1 alone and in complex with Ca2+ and an AMPA receptor tail peptide. BAR domains are banana-shaped homodimers that interact with cellular membranes and stabilize or sense membrane curvature. There is a direct correlation between the curvature of the BAR domain and the curvature of the membrane tubules that they stabilize. In the absence of diffracting crystals of the BAR domain of PICK1, we plan to use SAXS to determine the overall shape and radius of curvature of the BAR domain. By comparing the envelop of the BAR domain with that of full length PICK1 will also learn the location of the PDZ domain with respect to the BAR domain. In summary, the SAXS data will provide us a structural understanding of the mechanism of PICK1 activation by Ca2+ and the relative positions of the various domains of PICK1.

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。 与C-激酶1（PICK 1）相互作用的蛋白质是神经元中<$-氨基-3-羟基-5-甲基异恶唑-4-丙酸（AMPA）受体运输所必需的，AMPA受体运输是参与学习和记忆的关键过程。 PICK 1含有N-末端PSD-95/Discs-large/ZO-1（PDZ）结构域，随后是中央膜结合Bin/Amphiphysin/Rvs（BAR）结构域和C-末端酸性结构域。PICK 1受Ca 2+调节。在没有Ca 2+的情况下，PICK 1定位于细胞质，表明PICK 1通常是自抑制的。在Ca 2+刺激时，PICK 1通过其PDZ结构域结合AMPA受体尾，并通过其BAR结构域结合相邻的膜。 因此，一个主要的构象变化，预计将发生之间的Ca 2+结合和Ca 2+自由状态。 我们建议使用小角X射线散射（SAXS）来解决Ca 2+结合如何影响PICK 1的结构。 具体而言，我们计划确定PICK 1单独和与Ca 2+和AMPA受体尾肽复合的整体包膜。 BAR结构域是香蕉形的同源二聚体，其与细胞膜相互作用并稳定或感知膜曲率。 BAR结构域的曲率与它们稳定的膜小管的曲率之间存在直接相关性。在没有PICK 1的BAR域的衍射晶体的情况下，我们计划使用SAXS来确定BAR域的整体形状和曲率半径。通过将BAR域的包络与全长的包络进行比较，PICK 1还将获知PDZ域相对于BAR域的位置。总之，SAXS的数据将为我们提供一个结构的理解PICK 1激活的机制，由Ca 2+和PICK 1的各个结构域的相对位置。