Complexes of CAP in Transcription Activation

描述（由申请人提供）：我们目前的重点是对两个简单的三组分分子复合物的结构分析，这两个分子复合物是由E.卷曲分解代谢物激活蛋白（CAP）、RNA聚合酶（RNAP）和启动子DNA。CAP和RNAP及其与启动子DNA元件的单个复合物的结构知识是相当先进的，在过去几年中取得了特别迅速的进展。相比之下，关于CAP和RNAP在两个简单的转录激活类别中对启动子DNA的相互作用的知识仍然主要基于生物化学和遗传数据。拟议工作的目标是推进这些相互作用的结构知识，以产生完整的，准确的完整的转录复合物的分子图像。直接的目的是获得高分辨率的结构的子组件的I类和II类CAP-RNAP-启动子DNA复合物使用X-射线晶体学，和低分辨率的结构的完整的I类和II类CAP-RNAP-启动子DNA复合物使用冷冻电子显微镜。这项工作将产生完整的，准确的分子描述完整的转录起始复合物的两个主要的简单类CAP依赖性启动子，包括蛋白质-蛋白质相互作用，蛋白质-DNA相互作用，启动子DNA结构和溶剂的作用的描述。通过CAP的转录激活比细菌和真核生物中的大多数转录激活实例简单得多，所述细菌和真核生物中的转录激活可涉及大分子组分和/或DNA结合位点的复杂排列。CAP系统的拟议研究提供了一种手段，以迅速推进我们的理解的基本结构原则的转录控制。

DESCRIPTION (provided by applicant): Our current focus is on structural analysis of two simple three-component molecular complexes formed in transcription activation by the E. coil catabolite activator protein (CAP), RNA polymerase (RNAP), and promoter DNA. Structural knowledge of both CAP and RNAP and their individual complexes with promoter DNA elements is quite advanced, with particularly rapid progress made in the past few years. In contrast, knowledge about about the interactions between CAP and RNAP on promoter DNA in the two simple classes of transcription activation is still based largely on biochemical and genetic data. The goal of the proposed work is to advance structural knowledge of these interactions to yield complete, accurate molecular pictures of intact transcription complexes. The immediate aims are to obtain high-resolution structures of sub-assemblies of Class I and Class II CAP-RNAP-promoter DNA complexes using X-ray crystallography, and low resolution structures of intact Class I and Class II CAP-RNAP-promoter DNA complexes using cryo-electron microscopy. The work will yield complete, accurate molecular descriptions of intact transcription initiation complexes for the two major simple classes of CAP-dependent promoters, including descriptions of protein-protein interactions, protein-DNA interactions, promoter DNA structure, and role of solvent. Transcription activation by CAP is substantially simpler than most examples of transcription activation in bacteria and eukaryotes, which can involve complex arrangements of macromolecular components and/or DNA binding sites. The proposed studies of the CAP system provide a means to rapidly advance our understanding of fundamental structural principles underlying transcriptional control.