Dynamics of human Guanylate Binding Proteins: implications for their structure and oligomerisation

人鸟苷酸结合蛋白的动力学：对其结构和寡聚化的影响

• 批准号: 246179272
• 负责人: Professor Dr. Christian Herrmann
• 金额: --
• 依托单位: Lehrstuhl für Physikalische Chemie I
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2013
• 资助国家: 德国
• 起止时间: 2012-12-31 至 2019-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/246179272?language=en
• 关键词: Dynamics; human; Guanylate; Binding; Proteins

The ultimate goal of this project is to determine the supertertiary structure ensemble of human Guanylate binding Protein 1 (hGBP1) in solution and to unravel the relevance for its function. hGBP1 is a prototypic large GTP binding protein (GTPase). We are aiming for analysing its intra- and intermolecular protein dynamics in order to understand the molecular mechanism underlying membrane deformation and antimicrobial activity. Sophisticated single-molecule high-precision Förster Resonance Energy Transfer (FRET) experiments and species-specific fluorescence correlation spectroscopy (FCS) as well as kinetic and thermodynamic studies will be employed for defining precisely the architecture of the protein complexes in solution and the time courses of structural changes controlled by the nucleotides.The members of the protein family of large GTPases, like dynamin, Mx, and the numerous hGBP isoforms have many important biological functions in human cells, e.g. endocytosis and the defence of viral or microbrial attack by being part of inflamasomes. Despite these many distinct functions these proteins have many features in common such as the architecture of multiple similar domains, the nucleotide-dependent assembly in tetramers and the association larger oligomers to membranes. We will address many questions common to the members of the superfamily of large GTP binding proteins by pursuing our work on hGBP1 and stepwise including other GBP isoforms such as hGBP2, hGBP4 and hGBP5 (with 50-70% sequence identity but unknown structure) to unravel specific differences and similarities in the mechanism. By generating isoprenylated hGBP1, we also want to study how these structural changes are influenced by the association to membranes.Overall, we want to address the general question how the function of these large and flexible proteins is controlled by intramolecular domain and intermolecular subunit interactions, which modulate their overall supertertiary structure with distinct interface accessibility and dynamics. Especially we want to study how GTP binding and hydrolysis is exploited for function by modulating the molecular energy landscape and hence the dynamic equilibrium between thermodynamically stable conformational states. The detailed picture of a large GTPase, literally 'in action', will help us understanding on a molecular level how the nucleotide-driven protein-protein interactions can lead to concerted biological actions.

该项目的最终目标是确定溶液中人鸟苷酸结合蛋白1（hGBP 1）的超三级结构系综，并阐明其功能的相关性。hGBP 1是一种原型大GTP结合蛋白（GTbinding protein，GTbinding protein）。我们的目标是分析其内部和分子间的蛋白质动力学，以了解膜变形和抗菌活性的分子机制。先进的单分子高精度Förster共振能量转移（FRET）实验和物种特异性荧光相关光谱（FCS）以及动力学和热力学研究将用于精确定义溶液中蛋白质复合物的结构和核苷酸控制的结构变化的时间过程。并且众多的hGBP同种型在人细胞中具有许多重要的生物学功能，例如内吞作用和作为炎性体的一部分防御病毒或微生物攻击。尽管这些蛋白质具有许多不同的功能，但它们具有许多共同的特征，例如多个相似结构域的结构，四聚体中的核苷酸依赖性组装以及较大寡聚体与膜的缔合。我们将通过对hGBP 1的研究和逐步包括其他GBP亚型如hGBP 2，hGBP 4和hGBP 5（具有50-70%的序列同一性，但结构未知）来解决大GTP结合蛋白超家族成员共同的许多问题，以揭示机制中的特定差异和相似性。通过产生异戊二烯化的hGBP 1，我们还想研究这些结构变化是如何受到与膜的关联的影响的，总之，我们想解决这些大而灵活的蛋白质的功能是如何被分子内结构域和分子间亚基相互作用控制的一般问题，这些相互作用调节它们的整体超三级结构，具有不同的界面可及性和动力学。特别是，我们想研究如何利用GTP的结合和水解的功能，通过调节分子能量景观，从而在热力学稳定的构象状态之间的动态平衡。一个大的GTdR的详细图片，字面上的“在行动中”，将帮助我们在分子水平上理解核苷酸驱动的蛋白质-蛋白质相互作用如何导致协调一致的生物学行为。