FUNCTIONAL STUDY OF NATIVE AND SYNTHETIC NUCLEI

核孔复合体，一个100兆道尔顿的复合体，由 100个蛋白质的多个拷贝，是通过它的进口结构， 大量不同蛋白质和RNA底物的输出和循环 发生.该过程对约50 kDa以上的底物进行调节。这项建议 涉及脊椎动物NPC的结构和功能。第一个目标是 为了剖析NPC两个成员的分子相互作用和功能， Nup 98和Nup 153。这两种蛋白质都定位于NPC的核面; 有证据表明Nup 98与出口有关，Nup 153与进口有关。的位点 与靶向分子和NPC的其他成员的接触将被映射。 第二个目的是利用环状薄片，自发形成的结构 在卵细胞质中，类似于NPC，但更容易操纵。目标是 梳理这种复合物，以确定已知NPC的分子相互作用， 蛋白质和识别新的。环状薄片的形成 也可以在体外进行研究，这也将提供结构信息。在 最终目标私家侦探将利用一种新的检测方法， 标记配体，将标记的分子添加到环状薄片中， 构建反应，并鉴定了所涉及的标记蛋白。本试验将 可用于鉴定新的NPC蛋白，或测试与已知蛋白的相互作用。 全国人大选民。

The nuclear pore complex, a 100-megadalton complex comprised of 100 proteins in multiple copies, is the structure through which the import, export and cycling of a large number of diverse protein and RNA substrates occur. The process is regulated for substrates above~50 kDa. This proposal concerns the structure and function of the vertebrate NPC. The first aim seeks to dissect the molecular interactions and functions of two members of the NPC, Nup98 and Nup153. Both proteins are localized to the nuclear face of the NPC; evidence has linked Nup98 with export and Nup153 with import. The sites of contact with targeting molecules and other members of the NPC will be mapped. The second aim utilizes annulate lamellae, structures that form spontaneously in egg cytoplasm that resemble NPCs but are easier to manipulate. The goal is to tease apart this complex to identify molecular interactions of known NPC proteins and to identify new ones. The formation of the annulate lamellae will also be studied in vitro, which would also provide structural information. In the final aim, the P.I. will utilize a novel assay in which proteins bound to ligands are tagged, the tagged molecules are added to a annulate lamellae constitution reaction, and involved tagged proteins identified. This assay will be used to identify novel NPC proteins, or to test for interactions with known NPC constituents.