CAREER: Structural Basis of Transcription Termination in Thermophilic Eubacteria and Hypertext Curriculum for Advanced Undergraduate Biology Organized by Function and System

9734466弗里德曼该项目旨在开始阐明转录终止的结构基础。 转录终止似乎是由各种因子之间以及它们与RNA聚合酶之间的多种弱相互作用引起的。 已被鉴定为对正确转录终止至关重要的两种蛋白质是nusG和rho。 E.已知大肠杆菌rho是解旋酶，其在rho依赖性终止位点处将RNA转录物从转录泡解旋，而nusG充当rho和RNA聚合酶之间的衔接子。 从T.已经证明maritima容易通过蛋白水解被切割成两个片段。 两种碎片的结构将通过X射线晶体学确定。 同时，对T. maritima rho蛋白将由目前可获得的克隆构建。 还将进行该蛋白质的结晶实验，使用完整蛋白质和蛋白水解切割以分离RNA结合结构域的蛋白质。 最后，T.将研究maritima nusG和rho蛋白，以确定它们是否也显示出与在它们的E. coli对应物。 如果观察到这种相互作用，将启动对两种蛋白质复合物的结晶试验。 在这一职业建议的教育方面，将开发一套新的基于超文本的课程材料。 这些材料将被设计为以“垂直”方向呈现生物学，重点是生物功能或器官系统内不同组织水平之间的联系。 课程材料的纵向组织应有助于向本科生传达分子结构如何构成生物过程的基础。 它还应该为高年级的生物学本科生提供他们生物学知识的另一种观点，从而巩固他们在入门课上所学的知识。 这种研究和教育项目的结合将有助于进一步将结构生物学融入生物学的主流。 研究部分将带来一个新的生物学研究领域的结构调查，而教育部分将有助于我们目前的结构生物学知识整合到本科课程。

9734466 Friedman The project is designed to begin to elucidate the structural basis of transcription termination. Transcriptional termination appears to result from multiple weak interactions among various factors and between them and the RNA polymerase. Two proteins that have been identified as being critical for proper transcription termination are nusG and rho. E. coli rho is known to be a helicase that unwinds the RNA transcript from the transcription bubble at rho-dependent termination sites, while nusG acts as an adaptor between rho and the RNA polymerase. The nusG homolog from T. maritima has been demonstrated to be readily cleaved by proteolysis into two fragments. The structures of both fragments will be determined by x-ray crystallography. At the same time, expression systems for the T. maritima rho protein will be constructed from presently available clones. Crystallization experiments of this protein will also be conducted, both with the intact protein and protein proteolytically cleaved to separate the RNA-binding domain. Finally, potential interactions between the T. maritima nusG and rho proteins will be investigated to determine if they also show the same interaction which is seen in their E. coli counterparts. If this interaction is seen, crystallization trials on a complex of the two proteins will be initiated. On the education side of this CAREER proposal, a new set of hypertext-based curriculum materials will be developed. These materials will be designed to present biology in a "vertical" direction, focusing on the connections between different levels of organization within a biological function or organ system. The vertical organization of the curriculum material should help communicate to under-graduate students how molecular structure underlies biological processes. It should also provide upper-level biology undergraduates with an alternative view of their biological knowledge, thereby solidifying what they have been taught in their introductory classes. This combination of research and educational projects will help further integrate structural biology into the mainstream of biology. The research component will bring a new area of biological research under structural investigation, while the education component will help integrate our present knowledge of structural biology into the undergraduate curriculum.