Structure-function Studies of the Allosteric Mechanisms of Protein Switches

蛋白质开关变构机制的结构功能研究

• 批准号: 0919377
• 负责人: Marc Ostermeier
• 金额: $ 65.49万
• 依托单位: Johns Hopkins University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-01 至 2013-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=0919377&HistoricalAwards=false
• 关键词: Structure; function; Studies; Allosteric; Mechanisms

The activity of switch proteins can be turned on and off based on the binding of a signal molecule. Such effects are central to how cells carry out complex functions. Protein engineers are motivated to build switches of their own design to test their understanding of natural switches, to test hypotheses concerning the molecular basis of protein form and function, to create tools for elucidating cellular function and behavior, and to create switches for sensing and biomedical applications. The PI has created switches through fusing the genes encoding maltose binding protein (MBP) and TEM1 beta-lactamase (BLA). Understanding this switch's mechanism would inform the study of how such effects emerge through evolution and would facilitate future switch construction for applications. The objectives of this research effort are to develop an understanding of switch proteins as well as to establish design principles by which switches can be constructed. A structural model for MBP-BLA switch RG13 will be developed using a combination of NMR, computational modeling and crystallography. This model will be used to propose hypotheses on the switching mechanism and will then be tested experimentally using biophysical, biochemical and mutagenesis techniques.Broader ImpactsThe NMR and computational modeling work will provide tools for the study of large proteins. A senior undergraduate/graduate course combining computational protein design and directed evolution approaches will be co-taught and include a module on the protein switch work, in part to illustrate how interdisciplinary approaches are synergistic for addressing scientific problems. Graduate students supported by this grant would serve as teaching assistants in this course. Graduate students trained would have co-advisors from different scientific fields, receive interdisciplinary training and participate in meetings of professional societies. Undergraduates and high school students will participate in the planned research, increasing the three professors' participation in existing programs that provide research experience to high school students from a high school with a predominantly African American student body.

开关蛋白的活性可以基于信号分子的结合来开启和关闭。这些效应是细胞如何执行复杂功能的核心。 蛋白质工程师有动力构建自己设计的开关，以测试他们对天然开关的理解，测试有关蛋白质形式和功能的分子基础的假设，创建阐明细胞功能和行为的工具，并创建传感和生物医学应用的开关。 PI通过融合编码麦芽糖结合蛋白（MBP）和TEM 1 β-内酰胺酶（BLA）的基因创建了开关。了解这种开关的机制将有助于研究这种效应是如何通过进化出现的，并有助于未来的开关应用。这项研究工作的目标是发展开关蛋白的理解，以及建立开关可以构建的设计原则。MBP-BLA开关RG 13的结构模型将使用NMR、计算建模和晶体学的组合来开发。这个模型将被用来提出假设的开关机制，然后将实验测试使用生物物理，生物化学和诱变technology.Broader ImpactsThe核磁共振和计算建模工作将提供工具的研究大蛋白质。 一个高年级的本科生/研究生课程结合计算蛋白质设计和定向进化方法将共同授课，并包括蛋白质开关工作的模块，部分是为了说明跨学科的方法是如何协同解决科学问题。由该补助金资助的研究生将担任这门课程的助教。 接受培训的研究生将有来自不同科学领域的共同顾问，接受跨学科培训，并参加专业协会的会议。 本科生和高中生将参与计划中的研究，增加三位教授对现有项目的参与，这些项目为来自一所以非洲裔美国学生为主的高中的高中生提供研究经验。