CAREER: Dissecting Inter-domain Communication in Methionyl-tRNA Synthetase

该项目的最终目标是解释蛋白质- rna构象动力学。为了有效催化，来自大肠杆菌甲硫基tRNA合成酶（MetRS）反密码子结合域的信息必须在与该酶的同源tRNA相互作用时传递到远端活性位点。生化证据表明，这一信号事件需要RNA和蛋白质结构的改变。这种远距离的构象变化可能在核糖体和剪接体等大分子机器中非常常见，因此具有基本的生物学重要性，但它们实际上尚未被探索。适合于详细、定量分析的MetRS模型系统将有助于弥补这一差距。诱变、动力学和光谱学方法将用于研究MetRS结构域界面上的特定肽成分如何促进结构域间的通信。此外，晶体学将用于观察与同源tRNA和底物类似物的复合物以及与非配体酶的结构差异。这项跨学科研究的结果将有助于对氨基酰基- trna合成酶结构域间通讯的新理解，以及对具有远端配体结合和催化活性位点的酶的信号传导机制的一般认识。更广泛的影响：该项目将涉及维克森林大学生物学、化学和物理学领域的本科生和研究生教育。PI将她对科学研究的知识和热情带到了当地公立学校正在进行的推广项目中。这些教师讲习班和学生丰富活动的生物技术重点加强了当地的努力，以激励和培训农村和城市高中毕业生从事高等科学事业。这些努力既提高了私家侦探在研究和教育方面的职业发展，也提高了社会对经济增长的科学意识和培训。

This project's ultimate goal is to explain protein-RNA conformational dynamics.For efficient catalysis, information from the Escherichia coli methionyl-tRNA synthetase (MetRS) anticodon binding domain must be transmitted to a remote active site upon interaction with the enzyme's cognate tRNA. Biochemical evidence suggests this signaling event requires changes in both RNA and protein structure. Such long-range conformational changes are probably very common in macromolecular machines, such as ribosomes and spliceosomes, and, therefore, of fundamental biological importance, yet they remain virtually unexplored. The MetRS model system, amenable to detailed, quantitative analysis, will help to remedy this gap. Mutagenetic, kinetic, and spectroscopic approaches will be used to investigate how a specific peptide component at the domain interface of MetRS contributes to inter-domain communication. In addition, crystallography will be used to observe structural differences between MetRS in complex with its cognate tRNA and substrate analogs and with unliganded enyzme. Results obtained in this interdisciplinary study will contribute to a new understanding of inter-domain communication in aminoacyl-tRNA synthetases and to general knowledge about signaling mechanisms in enzymes with distant ligand-binding and catalytic active sites.Broader Impacts: The project will engage undergraduate and graduate education at the interface of biology, chemistry, and physics at Wake Forest University. The PI carries her knowledge of, and enthusiasm for, science research to ongoing outreach programs in local public schools. The biotechnology focus of these teacher workshops and student enrichment activities strengthens local efforts to motivate and to train rural and urban high school graduates to pursue postsecondary science careers. These efforts enhance both the PI's career development in research and education and the community's scientific awareness and training with an eye toward economic growth.