STRUCTURE OF AN ARCHAEAL TRANSCRIPTION COMPLEX

古细菌被认为是一种生命形式 区别于二十年前的细菌和桉树。的研究 古细菌转录装置表明，古细菌系统是 与真核生物中的系统高度同源。然而， 关于转录的详细机制或其在 古细菌。 本项目中的实验试图解决这些问题 通过定义转录的拓扑结构来回答问题 来自极端嗜热古菌激烈火球菌的复合体。 的 RNA聚合酶亚基相对于启动子DNA的排列， RNA转录本的转录水平将在 转录使用DNA-蛋白质和RNA-蛋白质光化学交叉， 链接方法 此外，国家行动方案与 转录起始因子TBP和TFB将被研究， 蛋白质-蛋白质足迹法和蛋白质交联法。 最后，一个假定的新转录因子的功能， 与TFB同源，将通过结合和 用多种激烈毕赤酵母启动子进行转录测定。 这些 实验应该提供一个很好的概述的结构， 古细菌转录装置，并将提供一个基础， 研究这一生物学领域的转录调控。 由于目前缺乏有关考古的基本知识， 生物学，这个提议中的实验很可能导致 发现并产生新的观察结果。 此外，同源性 真核生物和古细菌转录之间的差异意味着 从这些实验将直接适用于研究 真核转录 因此，这项工作可以提供健康 从人类遗传或获得的应用中产生的惠益 由基因转录异常引起的疾病。 以更好的 了解真核转录机制、杠杆点 可以识别和利用。

The archaea, or archaebacteria, were recognized as a form of life distinct from bacteria and eucarya just two decades ago. Studies of the archaeal transcription apparatus indicate that the archaeal system is highly homologous to the system in eucarya. However, little is known about the detailed mechanism of transcription or its regulation in the archaea. The experiments in this project attempt to address these questions by defining the topographical structure of the transcription complex from the hyperthermophilic archaeon Pyrococcus furiosus. The arrangement of the RNA polymerase subunits relative to promoter DNA and to the RNA transcript will be determined at defined steps in transcription using DNA-protein and RNA-protein photochemical cross- linking methods. In addition, the interactions between RNAP and the transcription initiation factors TBP and TFB will be investigated by protein-protein footprinting and protein cross-linking methods. Finally, the function of a putative novel transcription factor, homologous to TFB, will be investigated through binding and transcription assays with a variety of P. furiosus promoters. These experiments should provide an excellent overview of the structure of the archaeal transcription apparatus, and will provide a foundation for studying the regulation of transcription in this biological domain. Because of the current scarcity of basic knowledge concerning archaeal biology, the experiments in this proposal are very likely to result in discovery and to yield novel observations. In addition, the homology between archaeal and eucaryal transcription implies that conclusions from these experiments will be directly applicable to the study of eucaryal transcription. Therefore, this work may provide health benefits arising from applications to human inherited or acquired disease that originate from aberrant gene transcription. With a better understanding of eucaryal transcriptional mechanism, points of leverage for disease control may be identified and exploited.