CAREER: Defining the Chemical Basis of RNA Function by Nucleotide Analog Interference Mapping

Strobel教授的研究重点是建立一种新的方法来确定结构和催化重要RNA分子的结构-活性关系。目标分子包括一系列具有氧化不稳定的5'- o -（1-硫）三磷酸取代基的核苷酸类似物。单个类似物将通过体外转录并入RNA中，以创建突变RNA库，其中单个核苷酸被随机替换为类似物。在特定的生物系统中表现出活性的单个RNA突变体与那些没有活性的RNA突变体是不同的。随后在温和条件下用碘氧化裂解硫磷基团，并对测序凝胶上的活性和非活性类似物进行比较分析，提供了一种直接识别活性化学基团的方法。这种方法被称为核苷酸类似物干扰作图（NAIM），将用于鉴定RNA中与以下相关的化学基团：(1)5S rRNA与核糖体蛋白L25的相互作用；(2) U6 snRNA在pre-mRNA剪接中的作用；(3)神经孢子菌的自催化VS核酶；(4) α操纵子mRNA与S4蛋白的相互作用。教育活动包括开发生物有机课程，强调合成有机设计和大分子功能，并让当地本科生和少数民族学生参与研究项目。通过这个职业奖，有机动力学项目支持耶鲁大学分子生物物理和生物化学系的Scott Strobel教授的研究和教育活动。Strobel教授的研究重点是开发一种称为核苷酸类似物干扰映射的方法，该方法将大大加快识别结构和催化重要RNA分子中结构-活性关系的过程。Strobel教授的教学活动包括开发一个多部门的课程，探索使用有机化学合成来调查和理解生物系统，并通过让本地和少数民族本科生参与研究项目来促进对化学研究的兴趣。

Professor Strobel's research focuses on establishing a new methodology to determine structure-activity relationships in structurally and catalytically important RNA molecules. Target molecules include a series of nucleotide analogues with oxidatively labile 5'-O-(1-thio)triphosphate substituents. Individual analogues will be incorporated into RNA by in vitro transcription to create a library of mutant RNA's, where individual nucleotides have been randomly replaced by an analogue. Individual RNA mutants that exhibit activity in a particular biological system are distinguished from those that do not. Subsequent oxidative cleavage at the thiophosphate group under mild conditions with iodine, and comparative analysis of the active and nonactive analogues on sequencing gels offers a straightforward means to identify the active chemical group. This methodology, called Nucleotide Analogue Interference Mapping (NAIM), will be used to identify the chemical groups in RNA relating to: (1) the interaction of 5S rRNA with ribosomal protein L25; (2) the role of U6 snRNA in pre-mRNA splicing; (3) the autocatalytic VS ribozyme from Neurospora; and (4) the interaction of alpha operon mRNA with S4 protein. Educational activities include developing a bioorganic course emphasizing synthetic organic design and macromolecular function, and involving both local undergraduates and minority students in the research program. With this CAREER award, the Organic Dynamics Program supports the research and educational activities of Professor Scott Strobel of the Department of Molecular Biophysics and Biochemistry at Yale University. Professor Strobel's research is focused on developing a methodology, called Nucleotide Analogue Interference Mapping, that will significantly accelerate the process of identifying structure-activity relationships in structurally and catalytically important RNA molecules. Professor Strobel's educational activities include developing a multidepartmental course which will explore the use of organic chemical synthesis to investigate and understand biological systems, and promoting an interest in chemical research by involving both local and minority undergraduates in the research program.