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Structural and Functional Studies of Ubiquitin Binding Domains

泛素结合域的结构和功能研究

基本信息

批准号：
8349735
负责人：
James Hurley
金额：
$ 31.57万
依托单位：
NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
依托单位国家：
美国
项目类别：
财政年份：
资助国家：
美国
起止时间：
至
项目状态：
未结题

项目摘要

Structural and Functional Studies of Ubiquitin Binding Domains The covalent modification of proteins by ubiquitination is a major regulatory mechanism of protein degradation and quality control, endocytosis, vesicular trafficking, cell-cycle control, stress response, DNA repair, growth factor signaling, transcription, gene silencing, and other areas of biology. A class of specific ubiquitin binding domains mediates most of the effects of protein ubiquitination. The known membership of this group has expanded rapidly and now includes at least sixteen domains. The structures of many of the complexes with monoubiquitin have been determined, revealing interactions with multiple surfaces on ubiquitin. Inroads into understanding polyubiquitin specificity have been made for two UBA domains, whose structures have been characterized in complex with Lys48-linked diubiquitin. Several ubiquitin binding domains, including the UIM, CUE, VHS, and A20 ZnF, promote autoubiquitination, which regulates the activity of proteins that contain them. At least one of these domains, the A20 ZnF, acts as a ubiquitin ligase by recruiting a ubiquitin:ubiquitin conjugating enzyme thiolester adduct in a process that depends on the ubiquitin-binding activity of the A20 ZnF. The affinities of the monoubiquitin binding interactions of these domains span a wide range, but are most commonly weak, with Kd > 100 mM. The weak interactions between individual domains and monoubiquitin are leveraged into physiologically relevant high affinity interactions via several mechanisms: ubiquitin polymerization, modification multiplicity, oligomerization of ubiquitinated proteins and binding domain proteins, tandem binding domains, binding domains with multiple ubiquitin binding sites, and cooperativity between ubiquitin binding and binding through other domains to phospholipids and small G-proteins. The long terms goals of this project are to 1) determine the structural features of ubiquitin and its binding domains that are involved in molecular recognition; 2) correlate structural features with functional properties of these proteins in trafficking; and 3) understand the mechanisms whereby low-affinity interactions between individual binding domains and ubiquitin moieties and leveraged into physiological recognition events. One of the main questions remaining in the ESCRT field is how ubiquitinated cargo is handed off from ESCRT-0 coated domain into intralumenal buds, and to what extent deubiquitinating enzymes play an active role in this process. Another key goal is to understand the specificity of receptor ubiquitination upstream of the ESCRTs. Current efforts in the lab focus on these goals.

泛素结合结构域的结构与功能研究通过泛素化的蛋白质共价修饰是蛋白质降解和质量控制、胞吞作用、囊泡运输、细胞周期控制、应激反应、DNA修复、生长因子信号传导、转录、基因沉默和生物学的其他领域的主要调节机制。一类特异的泛素结合结构域介导蛋白质泛素化的大部分作用。这个组的已知成员迅速扩大，现在至少包括16个域。已经确定了许多与单泛素的复合物的结构，揭示了泛素与多个表面的相互作用。已经对两个乌巴结构域的多聚泛素特异性的理解取得了进展，其结构的特征在于与Lys 48连接的双泛素复合。几个泛素结合结构域，包括UIM、CUE、VHS和A20 ZnF，促进autoubiquitination，其调节含有它们的蛋白质的活性。这些结构域中的至少一个，A20 ZnF，通过在依赖于A20 ZnF的泛素结合活性的过程中募集泛素：泛素缀合酶硫酯加合物而充当泛素连接酶。这些结构域的monoubiquitin结合相互作用的亲和力跨越宽范围，但最常见的是弱的，Kd > 100 mM。通过几种机制，单个结构域和monoubiquitin之间的弱相互作用被利用成生理相关的高亲和力相互作用：泛素聚合，修饰多样性，泛素化蛋白和结合结构域蛋白的寡聚化，串联结合结构域，具有多个泛素结合位点的结合结构域，以及泛素结合和通过其它结构域与磷脂和小G蛋白结合之间的协同性。本项目的长期目标是：1）确定参与分子识别的泛素及其结合结构域的结构特征; 2）将这些蛋白质的结构特征与运输中的功能特性相关联; 3）了解单个结合结构域与泛素部分之间的低亲和力相互作用并利用到生理识别事件中的机制。 ESCRT领域中存在的主要问题之一是泛素化货物如何从ESCRT-0包被结构域传递到腔内芽中，以及去泛素化酶在此过程中发挥何种程度的积极作用。另一个关键目标是了解ESCRT上游受体泛素化的特异性。实验室目前的工作重点是这些目标。