Architecture of the Diffusion Barrier in the Nuclear Pore Complex

描述(申请人提供)：核孔复合体(NPC)是真核生物中最重要的分子机器之一，因为它门细胞的细胞质和核质之间的多孔管道，并控制所有的核质运输和通讯。它最重要的建筑特征是在其中心有一个鲜为人知的半渗透扩散屏障，该屏障对尺寸小至4纳米的细胞质蛋白质保持紧密的密封，但开放以便于传输尺寸高达40纳米的各种形状和大小的颗粒。这种灵活的屏障由一个名为FG核孔蛋白(FG NUP)的丝状蛋白家族组成，这些蛋白质以天然功能状态下的大未折叠结构域为特征，这些结构域装饰着多个苯丙氨酸甘氨酸基序(FG结构域)。我们提出的研究的具体目的是1)表征代表两种不同类型的FG NUP细丝的FG结构域的动态结构和分子内相互作用，这两种FG结构域锚定在NPC的三个不同的位置，2)验证这样的假设，即NUP的FG结构域之间的分子间相互作用在NPC中心创建了丝状网状结构，从而为蛋白质的被动扩散建立了大小选择性障碍。拟议的实验将结合生化、生物物理学、细胞生物学、结构(核磁共振)和分子建模技术，以深入了解FG NUP的动态行为、结构和功能。我们还将获得有关蛋白质无序结构域的动态行为和结构的基本知识。

DESCRIPTION (provided by applicant): The nuclear pore complex (NPC) is one of the most important molecular machines in eukaryotes because it gates the porous conduits between the cytoplasm and nucleoplasm of cells and controls all nucleo-cytoplasmic traffic and communication. Its most important architectural feature is a poorly understood semi-permeable diffusion barrier in its center that maintains a tight seal against cytoplasmic proteins as small as 4 nanometers in size, but opens to allow facilitated transport of particles of all shapes and sizes up to 40 nanometers in size. This flexible barrier is composed of a family of filamentous proteins named FG nucleoporins (FG nups) that feature large unfolded domains in their native functional state, which are decorated with multiple phenylalanine glycine motifs (FG domains). The specific aims of our proposed research are to 1) characterize the dynamic structure and intra-molecular interactions of FG domains representing two different types of FG nup filaments that are anchored at three different locales of the NPC, and 2) test the hypothesis that inter-molecular associations between FG domains of nups create a filamentous meshwork structure at the NPC center, which establishes the size-selective barrier to the passive diffusion of proteins. The proposed experiments will combine biochemical, biophysical, cell biological, structural (Nuclear Magnetic Resonance), and molecular modeling techniques to gain insight into the dynamic behavior, structure and function of the FG nups. We will also gain fundamental knowledge on the dynamic behavior and structure of disordered domains of proteins in general.