PECASE: A Difference Infrared Spectroscopic Study of a Nucleotide Binding Protein

9733566麦克唐纳这项职业计划整合了研究和教育部分，并涉及化学和生物专业的本科生以及高中教师在项目的各个方面。这项研究的目的是进一步了解参与DNA修复和重组的蛋白质的调节和机制。利用傅里叶变换红外(FTIR)差示光谱技术研究了大肠杆菌RecA蛋白的结构与功能之间的关系。这些研究应该会更好地理解RecA中负责调节DNA链交换的所有变构变化，这是一个对DNA重组和修复至关重要的过程。RecA的功能受核苷酸结合的调节。最初的研究将集中在三磷酸腺苷(ATP)和二磷酸腺苷(ADP)与RecA的结合。振动光谱将被用来识别ADP(与DNA亲和力降低的RecA构象)和ATP(与DNA亲和力较高的RecA构象)结合引起的蛋白质结构的特定变化。这些研究将在各种条件下进行，以调查核苷酸结合如何影响活性和非活性RecA核蛋白细丝中的蛋白质结构。这些研究将为未来的RecA研究提供基础，如(1)其他已知在体外与RecA相互作用的蛋白质如何影响先前研究的RecA结构变化，以及(2)实时跟踪核苷酸结合和链交换的时间分辨研究。高中教师，以及詹姆斯·麦迪逊大学化学和生物专业的本科生，以及中学教育辅修专业的学生，将在这项职业拨款的教育和研究方面发挥重要作用。更多的参与者可以通过提供两类项目(1)RecA的生化和差异红外研究(2)利用部门设备并可纳入新的以项目为导向的生物化学实验室的研究成果来参与研究工作。参与研究的学生将获得宝贵的研究经验，学习当前的生化和光谱技术，并对科学方法有更好的理解。本科生学员将有助于更好地理解RecA的结构-功能关系，以及一个令人兴奋的新生化实验室。将现在和未来的高中教师纳入这个项目最终将有助于吸引下一代科学家。

9733566 MacDonald This CAREER plan integrates research and educational components and involves undergraduate chemistry and biology majors as well as high school teachers in all aspects of the project. The goal of the research is to gain further insight into the regulations and mechanism of a protein involved in DNA repair and recombination. Fourier transform infrared (FTIR) difference spectroscopy will be used to investigate the relationship between protein structure and function in the Escherichia coli protein, RecA. These studies should result in a greater understanding of all allosteric changes in RecA that are responsible for regulating DNA strand exchange, a process crucial to DNA recombination and repair. RecA function is regulated by nucleotide binding. Initial studies will focus on the binding of adenosine triphosphate (ATP) and adenosine diphosphate (ADP) to RecA. Vibrational spectroscopy will be used to identify specific changes in protein structure induced by the binding of ADP (RecA conformation with decreased affinity for DNA), and ATP (RecA conformation with a higher affinity for DNA). These studies will be performed under a variety of conditions in order to investigate how nucleotide binding affects protein structure in both the active and inactive RecA nucleoprotein filaments. These studies will provide the groundwork for future RecA studies such as (1) how the previously studied changes in RecA structure are affected by other proteins known to interact with RecA in vitro and (2) time-resolved studies that follow nucleotide binding and strand exchange in real time. High school teachers, as well as James Madison University undergraduates in chemistry and biology ,and secondary education minors, will play instrumental roles in both the educational and research aspects of this CAREER grant. A larger number of participants can be involved in research efforts by providing two types of projects (1) biochemical and difference infrared studies of RecA (2) research effor ts that utilize departmental equipment and can be incorporated into the new project oriented biochemistry laboratory. Students involved will gain valuable research experience, learn current biochemical and spectroscopic techniques, and have a greater understanding of the scientific method. The undergraduate participants will contribute to a greater understanding of structure-function relationships in RecA and an exciting new biochemical laboratory. Including current and future high school teachers in this project will ultimately help to attract the next generation of scientists.