Crystallographic studies of retroviral intasome complexes

逆转录病毒嵌体复合物的晶体学研究

• 批准号: 8658560
• 负责人: Hideki Aihara
• 金额: $ 43.12万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-01 至 2018-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8658560
• 关键词: Acquired Immunodeficiency Syndrome; Address; Anti-HIV Agents; Antiviral Agents; Architecture; Binding; Biological Models; Cells; Chromatin; Chromosomes; Complex; Cryoelectron Microscopy; Crystallization; DNA; DNA Binding Domain; Detergents; Drug resistance; Enzymes; Genetic; Genome; Genomic DNA; Goals; HIV-1; Integrase; Integrase Inhibitors; Label; Left; Length; Light; Mercury; Methods; Molecular Chaperones; Mutagenesis; Mutation; Nucleoproteins; Phase; Protein Engineering; Proteins; Reaction; Research; Resolution; Retroviridae; Roentgen Rays; Rous sarcoma virus; Solubility; Spumavirus; Structure; Subfamily lentivirinae; System; Techniques; Testing; Vertebral column; Viral; Visna-maedi virus; X-Ray Crystallography; base; cellular targeting; clinically relevant; design; improved; inhibitor/antagonist; insight; lens epithelium-derived growth factor; next generation; novel strategies; particle; prototype; public health relevance; resistance mechanism; success; transcriptional coactivator p75; viral DNA

DESCRIPTION (provided by applicant): Retrovirus integration is catalyzed by the viral integrase (IN), which forms a tetrameric complex with both ends of the linear viral DNA genome and captures a cellular target DNA for concerted insertions of the viral DNA termini. The IN-DNA complex formed by HIV-1 IN is the target of a class of antiviral drugs called IN strand- transfer inhibitors. A better understanding of the IN-DNA interactions is therefore important for improving the clinically relevant anti-HIV drugs as well as designing next-generation IN inhibitors. The recent success in the structural studies of prototype foamy virus (PFV) IN-DNA complexes led to tremendous amount of insights into the retrovirus integration reaction. However, structural characterization of IN-DNA complexes from any other retrovirus systems by X-ray crystallography has remained elusive, leaving open the question as to how well the architecture of the IN-DNA complexes is conserved between distantly related retrovirus systems. We hypothesize that domain arrangement and IN-DNA interactions within the intasome, a nucleoprotein complex containing the tetramer of IN with the viral and target DNA molecules, are different between the canonical 3- domain IN including HIV-1 IN and the larger 4-domain PFV IN featuring an additional DNA-binding domain and longer inter-domain linkers. To address this hypothesis, we will determine crystal structures of the intasome complexes formed by the canonical 3-domain retroviral INs. Using a novel approach we have developed, crystals of the Rous sarcoma virus intasome suitable for structure determination have been obtained. We will further use our technique to pursue the crystal structure of a lentiviral intasome bound to the host co-factor LEDGF/p75. Our X-ray crystallographic analyses will provide the critically needed structural information to better understand the integration of HIV-1 and closely related retroviruses.

描述（申请人提供）：逆转录病毒整合由病毒整合酶（IN）催化，其与线性病毒DNA基因组的两端形成四聚体复合物，并捕获细胞靶DNA以协调插入病毒DNA末端。 HIV-1 IN 形成的 IN-DNA 复合物是一类称为 IN 链转移抑制剂的抗病毒药物的靶标。因此，更好地了解 IN-DNA 相互作用对于改进临床相关的抗 HIV 药物以及设计下一代 IN 抑制剂非常重要。最近原型泡沫病毒 (PFV) IN-DNA 复合物结构研究的成功使人们对逆转录病毒整合反应有了深入的了解。然而，通过X射线晶体学对任何其他逆转录病毒系统的IN-DNA复合物的结构表征仍然难以捉摸，这使得IN-DNA复合物的结构在远亲逆转录病毒系统之间的保守程度如何这一问题仍然悬而未决。我们假设，包含 HIV-1 IN 在内的规范 3 结构域 IN 和更大的 4 结构域之间，内体（一种包含 IN 四聚体与病毒和靶 DNA 分子的核蛋白复合物）内的结构域排列和 IN-DNA 相互作用是不同的PFV IN 具有额外的 DNA 结合域和更长的域间接头。为了解决这个假设，我们将确定由典型的 3 结构域逆转录病毒 IN 形成的嵌体复合物的晶体结构。使用我们开发的新方法，获得了适合结构测定的劳斯肉瘤病毒嵌体晶体。我们将进一步利用我们的技术来研究与宿主辅因子 LEDGF/p75 结合的慢病毒嵌体的晶体结构。我们的 X 射线晶体学分析将提供急需的结构信息，以更好地了解 HIV-1 和密切相关的逆转录病毒的整合。