Structural Studies of Nucleoprotein Complexes

核蛋白复合物的结构研究

• 批准号: 8013296
• 负责人: EDWARD H. EGELMAN
• 金额: $ 9.08万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-02-24 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8013296
• 关键词: Antibiotic Resistance; Area; BRCA2 gene; Bacteria; Bacteriophage P22; Binding; Binding Proteins; Biological; Biological Models; C-terminal; Cancer-Predisposing Gene; Capsid; Capsid Proteins; Collaborations; Complex; DNA; DNA Damage; Defect; Disease; Electron Microscopy; Environment; F-Actin; Filament; Gagging; Genetic Conjugation; Genetic Recombination; Grant; Herpesviridae; Herpesvirus 1; Heterogeneity; Higher Order Chromatin Structure; Homologous Gene; Human; Human Herpesvirus 4; Image Analysis; Infectious bursal disease virus; Inovirus; Kaposi Sarcoma; Knowledge; Lead; Light; Malignant Neoplasms; Methods; Murine leukemia virus; Mutate; N-terminal; Nucleoproteins; Pathway interactions; Pilum; Polymers; Population; Positioning Attribute; Property; Protein Binding; Protein Conformation; Protein Family; Proteins; Public Health; Regulation; Resolution; Role; Running; SS DNA BP; Site; Structure; Structure-Activity Relationship; System; Techniques; Tube; Viral; Viral Proteins; Virus; Virus Assembly; Work; genetic regulatory protein; innovation; insight; interest; leukemia; malignant breast neoplasm; polymerization; protein complex; public health relevance; tool

DESCRIPTION (provided by applicant): This long running project has been focused on nucleoprotein complexes active in DNA recombination and replication. The main tools that we have been using are electron microscopy and computational image analysis. Over the course of this project new and innovative methods have been developed that greatly increase our ability to understand the structure and function of many different helical polymers. What emerges in our studies of proteins like bacterial RecA and human Rad51 is that the polymers they form are highly dynamic, and that these polymers cannot be reduced to a single structure. Rather, the multiplicity of states that can exist needs to be understood. The relevance of these studies to disease has been greatly elevated by the understanding that proteins such as the product of the breast cancer susceptibility gene BRCA2 interact with Rad51 and target Rad51 to sites of DNA damage. In the proposed extension of this grant there are four aims: I) continued studies of complexes between RecA/Rad51 and the proteins that regulate these filaments, such as DinI, BRCA2, Hed1 and RAD51AP1. A focus will be on how the C-terminal domain of RecA and the N-terminal domain of Rad51 are regulatory, and how regulatory proteins that modulate the properties of these filaments interact with these regulatory domains. II) Determination of the structure of the filaments formed by viral single-stranded DNA binding proteins. The initial focus will be on Herpes Simplex Type 1 Virus (HSV-1) ICP8, but will then be extended to related viral proteins, such as from Epstein Barr Virus and Kaposi Sarcoma Herpes Virus. III) Structural studies of F-pili, needed for bacterial conjugation, and filamentous bacteriophage. It has long been suspected that the two are structural homologs, but structural studies of these filaments have been very difficult. We now have the tools in place to determine fairly high resolution structures for both. F-pili are involved in the transfer of antibiotic resistance within a bacterial population, so the public health relevance is great. IV) Capsid proteins that form spherical viral shells can be mutated or modified so that they form helical tubes instead. These tubes contain a conformation of the protein that fails to switch properly, and are thus of great interest in understanding virus assembly. The proposed focus is on capsid proteins from bacteriophage P22, Infectious Bursal Disease Virus, and Murine Leukemia virus. PUBLIC HEALTH RELEVANCE This project is aimed at understanding structure/function relationships in large complexes of proteins with DNA. Some of these proteins are active in genetic recombination, and defects in these recombination pathways can lead to cancer. Other complexes are involved in the transfer of antibiotic resistance within populations of bacteria, an area of growing concern for public health. And other complexes are involved in the assembly of viruses, including one that causes leukemia.

描述(由申请人提供)：这个长期运行的项目一直专注于活跃在DNA重组和复制中的核蛋白复合体。我们一直使用的主要工具是电子显微镜和计算图像分析。在这个项目的过程中，开发了新的和创新的方法，极大地提高了我们了解许多不同螺旋聚合物的结构和功能的能力。在我们对细菌RecA和人类RAD51等蛋白质的研究中发现，它们形成的聚合物是高度动态的，这些聚合物不能被还原为单一结构。相反，需要理解可能存在的多种国家。这些研究与疾病的相关性大大提高，因为人们了解到，乳腺癌易感基因BRCA2的产物等蛋白质与RAD51相互作用，并将RAD51靶向DNA损伤部位。在这笔拨款的拟议延期中，有四个目标：i)继续研究RecA/RAD51与调节这些细丝的蛋白质之间的复合体，如DINI、BRCA2、Hed1和RAD51AP1。重点将集中在RecA的C-末端结构域和RAD51的N-末端结构域是如何调节的，以及调节这些细丝特性的调节蛋白如何与这些调节结构域相互作用。Ii)确定病毒单链DNA结合蛋白形成的细丝的结构。最初的重点将是单纯疱疹病毒1型(HSV-1)ICP8，但随后将扩展到相关的病毒蛋白，如爱泼斯坦-巴尔病毒和卡波西肉瘤疱疹病毒。细菌接合所需的F-菌毛和丝状噬菌体的结构研究。长期以来，人们一直怀疑这两种纤维是结构上的同系物，但对这些细丝的结构研究一直非常困难。我们现在已经有了工具来确定这两种结构的相当高的分辨率。F-菌毛参与细菌种群内抗生素耐药性的转移，因此与公共卫生有很大的相关性。Iv)可以对形成球形病毒壳的衣壳蛋白进行突变或修饰，使其形成螺旋管。这些管子含有一种不能正确切换的蛋白质构象，因此对了解病毒组装非常有兴趣。建议的重点是噬菌体P22、传染性法氏囊病病毒和小鼠白血病病毒的衣壳蛋白。公共卫生相关性这个项目的目的是了解蛋白质与DNA的大型复合体中的结构/功能关系。其中一些蛋白质在基因重组中很活跃，这些重组途径中的缺陷可能会导致癌症。其他复合体还参与了细菌种群内抗生素耐药性的转移，这是一个日益受到公共卫生关注的领域。还有其他复合体参与病毒的组装，包括一种导致白血病的病毒。