Structure and mechanism of the ubiquitin activating enzyme

泛素激活酶的结构和机制

• 批准号: 191993457
• 负责人: Professor Dr. Hermann Schindelin
• 金额: --
• 依托单位: Rudolf-Virchow-Zentrum - Center for Integrative and Translational Bioimaging
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2010
• 资助国家: 德国
• 起止时间: 2009-12-31 至 2015-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/191993457?language=en
• 关键词: Structure; mechanism; ubiquitin; activating; enzyme

The activation of ubiquitin by the ubiquitin-activating enzyme (E1) represents the initial step in the covalent attachment of ubiquitin molecules to the side chains of lysine residues in target substrates. This modification is found in a multitude of cellular process including the selected degradation of the target protein by the proteasome. The E1-catalyzed activation of ubiquitin is a three-step process in which ubiquitin is adenylated at its C-terminal glycine, followed by the covalent attachment of ubiquitin to an essential cysteine residue. Finally, the enzyme ensures that ubiquitin is transferred in a trans-thioesterification reaction to one of several ubiquitin conjugating (E2) enzymes but not to any of the E2 enzymes of a ubiquitinlike protein modifier such as SUMO or NEDD8. Based on our crystal structure of the complex between the yeast E1 enzyme with ubiquitin bound at the adenylation active site, the first crystal structure of an intact ubiquitin E1, we intend to further characterize this enzyme with following aims in mind: (1) We will study how the enzyme recognizes cognate E2 enzymes and discriminates against E2 enzymes involved in SUMO or NEDD8 transfer. (2) We will characterize how the E1 enzyme is inhibited by small molecules to further probe its catalytic mechanism and as a first step towards the design of highly specific inhibitors. (3) We will determine the crystal structure of a quarternary complex consisting of the E1 enzyme, two bound ubiquitin molecules and an E2 enzyme to determine the conformational changes accompanying trans-thioesterification.

泛素活化酶（E1）对泛素的活化代表了泛素分子与靶底物中赖氨酸残基侧链共价连接的初始步骤。这种修饰存在于许多细胞过程中，包括蛋白酶体对靶蛋白的选择性降解。E1催化的泛素活化是一个三步过程，其中泛素在其C-末端甘氨酸被腺苷酸化，然后泛素共价连接到必需的半胱氨酸残基。最后，该酶确保泛素在硫酯交换反应中转移到几种泛素缀合（E2）酶中的一种，但不转移到泛素样蛋白修饰剂如SUMO或NEDD 8的任何E2酶。基于我们的酵母E1酶与泛素结合在腺苷酸化活性位点之间的复合物的晶体结构，完整的泛素E1的第一个晶体结构，我们打算进一步表征这种酶，并考虑到以下目标：（1）我们将研究该酶如何识别同源E2酶并区分参与SUMO或NEDD 8转移的E2酶。(2)我们将描述E1酶如何被小分子抑制，以进一步探索其催化机制，并作为设计高度特异性抑制剂的第一步。(3)我们将确定由E1酶，两个结合的泛素分子和E2酶组成的四元复合物的晶体结构，以确定伴随着转硫酯化的构象变化。