Expanding Mechanistic Insights into Protein Ubiquitylation

扩展对蛋白质泛素化的机制见解

• 批准号: 10330645
• 负责人: Rachel E Klevit
• 金额: $ 68.76万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-08-15 至 2027-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10330645
• 关键词: Address; Affect; BARD1 gene; BRCA1 gene; Biochemical; Biochemical Reaction; Biophysics; Cell physiology; Cells; DNA Damage; Data; Development; Enzymes; Family; Functional disorder; Genetic Transcription; Human; Human Genome; Knowledge; Lysine; Modification; Process; Protein Engineering; Proteins; Reaction; Research; Site; Structure; System; Translations; Tumor Suppressor Proteins; Ubiquitin; Work; cell growth regulation; human disease; insight; malignant breast neoplasm; member; protein function; response; stem; therapeutic target; ubiquitin-protein ligase

PROJECT SUMMARY Covalent attachment of ubiquitin (Ub) to other proteins is among the most widespread and diverse modes of eukaryotic cellular regulation. The modification occurs on practically every protein in a cell at some point in its lifetime and is itself highly diverse. The type of ubiquitylation determines a product’s fate and a protein may undergo different modes of ubiquitylation depending on cellular circumstances. The origins of this diversity stem from the protein machinery responsible for Ub attachment. A trio of enzymes, E1, E2, and E3 coordinate the process, with several E1s, dozens of E2s, and many hundreds of E3s encoded in the human genome. Over the past 20 years, we have asked fundamental questions about how E2s and E3s work and have contributed to the structural, biochemical, and mechanistic understandings of the field. Our work began with the breast cancer tumor suppressor, BRCA1/BARD1 that was among the earliest RING-type E3 ligases to be identified. Over the years, we have expanded to study numerous E3s and E2s, making many unexpected discoveries along the way. The wide reach of protein ubiquitylation in cellular function means that dysfunction of components is associated with myriad human diseases and developmental issues. Such associations make the Ub system attractive for therapeutic targeting. Direct targeting of the ubiquitylation machinery as well as efforts to re-engineer protein ubiquitylation machinery to selectively target a specific cellular protein are both proving to be powerful strategies. Such translational efforts rely implicitly on mechanistic understanding and reveal the power of well- grounded structure/function research. Despite the apparent maturity of the field, there is still much we do not understand at a fundamental level. We do not know the full range of biochemical reactions carried out by the ~30 human E2s as fully one-quarter of these are uncharacterized. Existing data reveal that not all E2s carry out the presumed reaction that attaches Ub to lysine sidechains, implying the existence of ubiquitylated species that have yet to be investigated in cells. Second, understanding of how E2/E3s carry out mono-ubiquitylation is lacking. Unlike poly-ubiquitylation, attachment of a single Ub (mono-Ub) tends to occur in a site- selective manner implying that substrates to be mono-ubiquitylated are handled differently from those destined to have chains built upon them. Third, lack of knowledge regarding how mono- Ub attachment affects the structure and function of proteins limits understanding of how the modification regulates critical cellular processes including transcription, translation, and DNA damage response, among others.

项目摘要 泛素（UB）与其他蛋白质的共价附着是最广泛的蛋白质之一 真核细胞调节的潜水模式。修改实际上发生 细胞中的蛋白质在其一生的某个时候，本身是高度潜水员。泛素化的类型 确定产品的命运，蛋白质可能会经历不同的泛素化模式 取决于细胞情况。这种多样性的起源源于蛋白质 负责UB附件的机械。三个酶，E1，E2和E3协调 过程，有几个E1，数十个E2和人类编码的数百个E3 基因组。在过去的20年中，我们询问了有关E2和如何方式的基本问题 E3工作，并为结构，生化和机械理解做出了贡献 田地。我们的工作始于乳腺癌肿瘤抑制剂BRCA1/BARD1 在最早的环型E3连接酶中。多年来，我们已经扩展到 研究许多E3和E2，在此过程中引起了许多意外的发现。 细胞功能中蛋白质泛素化的广泛范围是指 组成部分与无数的人类疾病和发育问题有关。这样的 关联使UB系统对治疗靶向具有吸引力。直接定位 泛素化机制以及重新设计蛋白质泛素化机械的努力 有选择地靶向特定的细胞蛋白都被证明是强大的策略。 翻译努力隐含地依赖于机械理解并揭示了良好的力量 接地结构/功能研究。尽管该领域显然成熟，但仍然存在 我们在基本层面上不了解。我们不知道全部 〜30人E2进行的生化反应是完全四分之一的 未表征。现有数据表明，并非所有E2都会执行提出的反应 将UB附在赖氨酸侧chains上 在细胞中进行研究。其次，了解E2/E3如何进行单次启发化是 缺乏。与多泛素化不同，单个UB（单ub）的附着倾向于在位点发生 选择性的方式表明，要单次泛素化的底物的处理方式不同 那些注定要建立连锁店的人。第三，缺乏有关单一的知识 UB附件影响蛋白质的结构和功能，限制了对 修改调节关键细胞过程，包括转录，翻译和DNA 损坏反应等。