CAREER: Biochemical and Structural Mechanisms Controlling tRNA-Modifying Metalloenzymes

职业：控制 tRNA 修饰金属酶的生化和结构机制

• 批准号: 2339759
• 负责人: Jeffrey Mugridge
• 金额: $ 82万
• 依托单位: University of Delaware
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2024
• 资助国家: 美国
• 起止时间: 2024-03-01 至 2029-02-28
• 项目状态: 未结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=2339759&HistoricalAwards=false
• 关键词: CAREER; Biochemical; Structural; Mechanisms; Controlling

With the support of the Chemistry of Life Processes (CLP) program in the Division of Chemistry (CHE) and the Established Program to Stimulate Competitive Research (EPSCoR), Jeffrey Mugridge from the University of Delaware is studying the biochemical and structural mechanisms controlling metalloenzymes that install important chemical modifications on transfer RNA (tRNA) molecules. Cellular tRNA molecules are decorated with a huge diversity of chemical modifications that are essential for correctly tuning tRNA structure, stability, and efficient and accurate protein synthesis. Defects in the enzymes and biochemical pathways that install tRNA modifications are linked to a wide range of human diseases from cancers to neurodegenerative disorders. However, for many of the enzymes that install key chemical modifications on tRNA, a clear understanding of how these enzymes selectively carry out their modification reactions is missing. Filling this gap in knowledge is important for our fundamental understanding of cellular RNA biology. This proposal will define the detailed atomic-level mechanisms for two distinct classes of metal-dependent enzymes (metalloenzymes) that install key modifications on tRNA that directly impact protein synthesis. Closely integrated with this research, a course-based undergraduate research experience (CURE) will be developed that engages both undergraduates and high-school students from underrepresented groups in a collaborative, discovery-based course where students will make and experimentally test predictions about protein-tRNA interactions. The outcomes from this work could provide new information on how tRNA-modifying metalloenzymes carry out complex, multistep reactions on tRNA and broaden engagement of students in research-focused activities at the undergraduate and high-school levels.Hypermodification of the tRNA anticodon loop is essential for proper codon-anticodon recognition and mRNA decoding in the ribosome during translation. The cell installs numerous, chemically complex modifications at these locations to ensure and control translational efficiency and fidelity. This project combines techniques from biochemistry, structural biology, biophysics, and chemical biology to study two different classes of tRNA-modifying metalloenzymes that install sequential modifications on the anticodon loop of tRNA. The goals of this work are to: (1) define the stepwise chemical mechanisms used by these classes of metalloenzymes to install anticodon loop modifications, (2) unveil the structural mechanisms and protein conformations that control tRNA recognition and modification selectivity, and (3) provide new, broad information about how these classes of metalloenzymes are regulated in the cell. The research outcomes could deepen our basic understanding of tRNA biology and pave the way for future therapeutic development targeting these or similar metalloenzymes and tRNA modification pathways.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

在化学系（CHE）生命过程化学（CLP）项目和刺激竞争研究既定项目（EPSCoR）的支持下，特拉华大学的Jeffrey Mugridge正在研究控制金属酶的生化和结构机制，金属酶在转移RNA （tRNA）分子上安装重要的化学修饰。细胞tRNA分子被大量的化学修饰修饰，这些修饰对于正确调整tRNA结构、稳定性和高效准确的蛋白质合成至关重要。安装tRNA修饰的酶和生化途径中的缺陷与从癌症到神经退行性疾病的广泛人类疾病有关。然而，对于许多在tRNA上安装关键化学修饰的酶来说，对这些酶如何选择性地进行修饰反应的清楚理解是缺失的。填补这一知识空白对于我们对细胞RNA生物学的基本理解非常重要。该提案将定义两种不同类型的金属依赖性酶（金属酶）的详细原子水平机制，它们在tRNA上安装直接影响蛋白质合成的关键修饰。与本研究紧密结合，将开发一种基于课程的本科生研究体验（CURE），吸引来自代表性不足群体的本科生和高中生参加一门合作的、基于发现的课程，学生们将对蛋白质- trna相互作用进行预测并进行实验测试。这项工作的结果可以为tRNA修饰金属酶如何在tRNA上进行复杂的多步骤反应提供新的信息，并扩大学生在本科和高中阶段的研究重点活动的参与。tRNA反密码子环的超修饰是翻译过程中核糖体正确识别密码子-反密码子和mRNA解码所必需的。细胞在这些位置安装大量的化学复杂修饰，以确保和控制翻译效率和保真度。本项目结合了生物化学、结构生物学、生物物理学和化学生物学的技术，研究了两种不同类型的tRNA修饰金属酶，它们在tRNA的反密码子环上进行了顺序修饰。这项工作的目标是：(1)定义这些金属酶用于安装反密码子环修饰的逐步化学机制，(2)揭示控制tRNA识别和修饰选择性的结构机制和蛋白质构象，以及(3)提供关于这些金属酶如何在细胞中被调节的新的，广泛的信息。该研究结果可以加深我们对tRNA生物学的基本认识，并为未来针对这些或类似的金属酶和tRNA修饰途径的治疗开发铺平道路。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。