STRUCTURAL STUDY OF THE OUTER SHELL OF THE COPII VESICLE COAT

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。这是旨在获得酵母COPII囊壳结构信息的一系列建议中的第三项。COPII是一个420 kDa的五聚体复合体，负责从内质网(ER)萌发运输小泡，并将特定的货物内容物输送到高尔基复合体。在第一个提案中，我们重点研究了COPII涂层的内壳，Sec23/24-Sar1复合体。我们能够确定SEC24与货物和陷阱序列的三个复合体的结构(发表在《细胞》杂志上的论文)。在目前的提案中，我们把重点放在COPII的外壳上，它由Sec13/31复合体组成。SEC 13/31在功能上与笼状蛋白同源，但在结构上与笼状蛋白同源--它使内质网膜上的COPII内壳聚合，导致膜变形。由于像笼状蛋白一样，Sec13/31是一种能够齐聚的长而灵活的结构(如EM图像所示)，因此完整的复合体不太可能被结晶。因此，我们已经将蛋白质解剖成我们认为是两个功能感兴趣的片段，这构成了我们在本申请中提出的两个项目。

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. This is the third in a series of proposals that aims at obtaining structural information on the yeast COPII vesicalar coat. COPII is a 420 kDa pentameric complex responsible for budding transport vesicles from the endoplasmic reticulum (ER) and delivering the specific cargo contents to the Golgi complex. In the first proposal we focused on the inner shell of the COPII coat, the Sec23/24-Sar1 complex. We were able to determine the structures of three complexes of Sec24 bond to cargo and SNARE sequences (paper published in Cell). In the current proposal we focus on the outer shell of COPII , comprising of the Sec13/31 complex. Sec 13/31 is functionally, but not structurally, homolosous to clathrin--it polymerizes the COPII inner shell on the ER membrane and causes membrane deformation. Since, like clathrin, Sec13/31 is a long, flexible structure capable of oligomerizing (as seen in EM images) it is unlikely that the intact complex can be crystallized. There fore, we have dessected the protein into waht we believe are the tow functionally interesting fragments, and these constitue the two projects that we propose in this application.