HIV-1 Integrase Structural Biology

HIV-1 整合酶结构生物学

• 批准号: 7905212
• 负责人: Alan N. Engelman
• 金额: $ 5.64万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-12 至 2010-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7905212
• 关键词: Arts; Binding; Binding Sites; Biochemical; Biological Assay; C-terminal; Catalytic Domain; Cells; Chromosomes; Clinical Trials; Complementary DNA; Complex; Crystallization; Crystallography; DNA; Data; Drug Delivery Systems; Drug Design; Enzymes; Equilibrium Centrifugation; Gel Chromatography; Glean; HIV-1; HIV-1 integrase; Hot Spot; Housing; Human; In Vitro; Integrase; Integrase Inhibitors; Isotonic Exercise; Length; Molecular; Molecular Weight; Mutation; N-terminal; NMR Spectroscopy; Pathway interactions; Pattern; Phase; Proteins; Reaction; Retroviridae; Reverse Transcription; Site; Sodium Chloride; Solubility; Source; Structure; Suicide; System; Techniques; Viral; Work; base; design; dimer; enzyme structure; inhibitor/antagonist; killings; lens epithelium-derived growth factor; light scattering; mutant; novel; research study; sedimentation equilibrium; small molecule; structural biology; synchrotron radiation; three dimensional structure; transcriptional coactivator p75

Like all retroviruses, HIV-1 must integrate the cDNA copy made by reverse transcription into a cell chromosome in order to replicate. HIV-1integration is catalyzed by the essential viral enzyme integrase. This makes the integrase an attractive drug target, and anti-integrase compounds are in US clinical trials. Rational drug design benefits from visualizing three-dimensional enzyme structures, and numerous structures of the HIV-1 integrase catalytic core domain have been solved. Although two domain N-terminal/core and core/C-terminal structures are also solved, there is no structure for the intact integrase protein, which in large part can be attributed to poor protein solubility. In cells, integrase is likely to interact with normal human proteins, and the interaction between integrase and the transcriptional co-activator lens epithelium-derived growth factor (LEDGF) forms the basis for this application. We recently solved the three-dimensional structure of the integrase-binding domain (IBD)in LEDGF using NMR spectroscopy and herein present the crystal structure of the IBD bound to the dimeric core domain of HIV-1 integrase. Building from these results, we propose to utilize the LEDGF IBD to form novel LEDGF-IN complexes and to solve the three-dimensional structure of the full-length integrase protein. Our novel structure revealed a pocket at the core domain dimer interface that is occupied by LEDGF hot spot residues upon complex formation. Because integrase mutations that ablate the interaction with LEDGF kill HIV-1, we hypothesize that compounds that bind to this region of the core and preclude LEDGF binding might similarly cripple HIV-1. Previous results revealed that certain integrase inhibitors bind at or near this pocket, but those compounds did not inhibit the LEDGF-integrase interaction. A novel assay system will be designed to select for inhibitors of the LEDGF-integrase interaction. The results of these experiments will significantly aid the design of inhibitors of HIV-1 integrase function, as the three-dimensional structure of the intact enzyme will be elucidated and an assay that could select for novel inhibitors of HIV-1 replication will be developed.

与所有逆转录病毒一样，HIV-1 必须将逆转录产生的 cDNA 拷贝整合到细胞中 染色体以进行复制。 HIV-1 整合由必需病毒酶催化 整合酶。这使得整合酶成为有吸引力的药物靶点，并且抗整合酶化合物正在 美国临床试验。合理的药物设计受益于三维酶的可视化 HIV-1整合酶催化核心结构域的许多结构已经得到解决。 虽然也解决了两个域N端/核心和核心/C端结构，但没有 完整整合酶蛋白的结构，这在很大程度上可归因于较差的蛋白质 溶解度。在细胞中，整合酶很可能与正常的人类蛋白质相互作用，并且这种相互作用 整合酶和转录共激活因子晶状体上皮衍生生长因子之间 (LEDGF) 构成了该应用的基础。我们最近解决了三维结构 使用 NMR 光谱分析 LEDGF 中的整合酶结合域 (IBD)，并在此介绍 与 HIV-1 整合酶二聚体核心结构域结合的 IBD 的晶体结构。建筑自 根据这些结果，我们建议利用 LEDGF IBD 形成新型 LEDGF-IN 复合物并 解析全长整合酶蛋白的三维结构。我们新颖的结构 揭示了核心域二聚体界面处的一个口袋，该口袋被 LEDGF 热点残基占据 复杂形成后。因为整合酶突变会消除与 LEDGF 的相互作用，从而杀死 HIV-1，我们假设与核心的该区域结合并排除 LEDGF 的化合物 结合可能同样会削弱 HIV-1。先前的结果表明某些整合酶抑制剂结合 在该口袋处或附近，但这些化合物不会抑制 LEDGF-整合酶相互作用。一本小说 检测系统将被设计为选择 LEDGF-整合酶相互作用的抑制剂。这 这些实验的结果将极大地帮助设计 HIV-1 整合酶功能的抑制剂， 因为完整酶的三维结构将被阐明，并且可以进行测定 将开发新的 HIV-1 复制抑制剂。