Expanding Mechanistic Insights into Protein Ubiquitylation

扩展对蛋白质泛素化的机制见解

• 批准号: 10796652
• 负责人: Rachel E Klevit
• 金额: $ 9.52万
• 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/10796652
• 关键词: 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连接是最广泛和最重要的。 真核细胞调节的不同模式。几乎每一次修改都发生在 蛋白质在其生命周期中的某个时刻存在于细胞中，并且本身是高度多样的。泛素化的类型 决定了产物的命运，蛋白质可能经历不同的泛素化模式 取决于细胞环境。这种多样性的起源源于蛋白质 负责UB附件的机械。三种酶E1、E2和E3负责协调 过程中，有几个E1，几十个E2，和数百个E3编码在人类 基因组在过去的20年里，我们一直在问E2和 E3的工作，并作出了贡献的结构，生物化学和机械的理解 的领域。我们的工作始于乳腺癌肿瘤抑制因子BRCA 1/BARD 1， 是最早被鉴定的RING型E3连接酶。多年来，我们已扩大到 研究了大量的E3和E2，在研究过程中有许多沿着意想不到的发现。 蛋白质泛素化在细胞功能中的广泛影响意味着， 成分与无数人类疾病和发育问题有关。等 关联使得Ub系统对于治疗靶向具有吸引力。直接针对 泛素化机制以及努力重新设计蛋白质泛素化机制， 选择性地靶向特定的细胞蛋白质都被证明是强大的策略。等 翻译工作隐含地依赖于机械的理解，并揭示了良好的力量， 结构/功能研究。尽管该领域已明显成熟，但仍有 很多我们在基本层面上不理解的东西。我们并不知道 大约30个人类E2进行的生化反应，其中整整四分之一是 没有特征的现有的数据显示，并非所有的E2都能进行假定的反应， 将Ub连接到赖氨酸侧链上，这意味着存在泛素化的物种， 在细胞中进行研究。第二，理解E2/E3如何进行单泛素化， 缺乏与多聚泛素化不同，单个Ub（mono-Ub）的连接往往发生在一个位点， 选择性方式意味着要被单泛素化的底物的处理不同于 那些注定要被锁链束缚的人第三，缺乏对单方面知识的了解， UB连接影响蛋白质的结构和功能，限制了对蛋白质结构和功能的理解。 修饰调节关键的细胞过程，包括转录，翻译和DNA 损害赔偿等。