Structural-Function Studies of Shared hRNAP Components

描述(申请人提供)：本项目的长期目标是 了解AGEN转录调控的机制 在分子水平上的表达。所有真核细胞的转录 基因由三个不同的RNI聚合酶(RPS)I、II和III执行， 它们分别合成核糖体、信使和转移RNA。RP是 多亚单位复合体，由酶特异性多肽和 三种酶共有的亚基。人类共同亚基(hRPB5，hRPB6， HRPB8、hRPB10-α和hRPB10-β)是真正的通用转录 因为它们对所有人类基因的表达都是必不可少的。 三个人力资源主管。因此，确定这些亚基的结构和功能 是阐明基本的分子机制的关键一步 控制全球人类基因转录。首席调查员已经 用核磁共振确定hRPB6的溶液结构 (核磁共振)光谱学。本提案侧重于结构分析 人类常见亚基及其在分子中的作用 人类RPII的结构和功能。具体目标和实验 所提出的研究方法是：1)进行高分辨率的 共同亚基hRPB8，hRPB10-α，hRPB10-β， 采用X-射线结晶学和核磁共振波谱。2)确定 HRP共同亚基与一般亚基相互作用的结构基础 转录起始和延伸因子。在这一目标中，建议 对hRPB5亚基进行X射线结晶学分析 转录起始因子hTFIIB和hRPB6亚单位与 转录延伸因子hTFIIS。3)剖析 肿瘤功能中常见的亚基hRPB10a、hRPB10beta和hRPB8 抑制蛋白。 这些研究将提供高分辨率的三维结构 人类共同的亚基，并将阐明它们在组装中的作用， 三个核hrp的结构和功能。的原子坐标 常见的亚基，除了它们的科学价值外，还可以用于 基于结构的合理设计的化学品，可以选择性地改变 HRPs的结构和功能及其在分子领域的潜在应用 医药。

DESCRIPTION (provided by applicant): The long-term objective of this project is to understand the mechanisms of the transcriptional regulation of gent expression at the molecular level. Transcription of all eukaryotic nuclear genes is performed by three distinct RNI polymerases (RPs) I, II, and III, which synthesize ribosomal, messenger, and transfer RNAs, respectively. RPs are multi-subunit complexes, consisting of enzyme-specific polypeptides and subunits common to all three enzymes. The human common subunits (hRPB5, hRPB6, hRPB8, hRPB10-alpha, and hRPB10-beta) are bona fide universal transcription factors because they are essential for the expression of all human genes by the three hRPs. Therefore, determining the structure and function of these subunits is an essential step towards elucidating the basic molecular mechanisms that control global human gene transcription. The principal investigator has already determined the solution structure o hRPB6 using Nuclear Magnetic Resonance (NMR) spectroscopy. The present proposal focuses on the structural analysis of the common human subunits and the elucidation of their roles in the molecular architecture and function of human RPII. The specific aims and experimental approaches of the proposed research are: 1) To perform a high-resolution structural analysis of the common subunits hRPB8, hRPB10-alpha, hRPB10-beta, using X-ray crystallography and NMR spectroscopy. 2) To determine the structural basis for the interactions between hRP common subunits and general transcription initiation and elongation factors. In this aim it is proposed to perform an X-ray crystallographic analysis of the hRPB5 subunit in complex with transcription initiation factor hTFIIB and of the hRPB6 subunit in complex with the transcription elongation factor hTFIIS. 3) To dissect the role of the common subunits hRPB10alpha, hRPB10beta, and hRPB8 in the function of tumor suppressor proteins. These studies will provide high-resolution three-dimensional structures of the human common subunits and will elucidate their roles in the assembly, structure, and function of the three nuclear hRPs. The atomic coordinates of the common subunits, besides their scientific value, could also be used for structure-based rational design of chemicals that could modify selectively the structure and function of the hRPs, with potential applications in molecular medicine.