Structural Basis of Antitermination by the E. coli Phages lambda and HK022

目前的资助申请描述了x射线晶体学，核磁共振光谱和分子动力学计算，光谱学和生化方法相结合的独特努力，以阐明大肠杆菌（E. coli）中具有真核生物类似物的非常基本的转录调节过程的结构基础。转录终止和反终止的调控是大量病毒复制的重要调控机制，包括原型大肠杆菌噬菌体¿，大肠杆菌噬菌体香港022 (HK022)，真核免疫缺陷病毒如人类免疫缺陷病毒1 （HIV-1）以及许多其他病毒。目前的应用是针对噬菌体¿/大肠杆菌和HK022/大肠杆菌的终止/抗终止系统的三维结构。长期目标是扩大我们对原核生物中抗终止转录调控基础的认识，最终目标是合理设计新的抗生素，并为合理设计针对慢病毒的药物提供基础。特别是，我们建议确定完整噬菌体¿/大肠杆菌抗终止体和噬菌体HK022/大肠杆菌终止复合物的主要三维结构部分。除了噬菌体编码的抗终止蛋白¿N和Nun以及调控RNA元件外，这些复合物中的关键调控蛋白是NusA和NusG，我们的研究重点是NusA和NusG。

The current grant application describes a unique effort towards the combination of X-ray crystallography, NMR-spectroscopy and molecular dynamics calculations, optical spectroscopy and biochemical methods to shed light onto the structural basis of a very fundamental transcription regulatory process in the bacterium Escherichia coli (E. coli) that has analogs in eukaryotes. Modulation of transcription termination and antitermination is an essential regulatory mechanism of the replication of a large number of viruses including the prototypical E. coli phage ¿, the E. coli phage Hongkong 022 (HK022), the eukaryotic immunodeficiency viruses such as human immunodeficiency virus 1 (HIV-1) as well as a number of other viruses. The current application targets the three-dimensional structure of the termination/ antitermination systems of phage ¿/ E. coli and of HK022/ E. coli. The long-term goal is to extend our knowledge of the basis of transcription regulation by antitermination in prokaryotes, with the eventual goal of rationally designing new antibiotics, and to provide a basis for the rational design of drugs against lentiviruses. In particular, we propose to determine major parts of the three-dimensional structure of the complete phage ¿/ E. coli antitermination and the phage HK022/ E. coli termination complex. Besides phage-encoded antitermination proteins ¿N and Nun, respectively, and regulatory RNA elements, the key regulator proteins in these complexes are NusA and NusG, on which our studies are focused on.