Biochemical Mechanism of HIV DNA Integration

HIV DNA整合的生化机制

• 批准号: 7867673
• 负责人: Alan N. Engelman
• 金额: $ 47.5万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-07-01 至 2015-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7867673
• 关键词: AT-Hook Motifs; Adverse effects; Antiviral Therapy; Binding; Binding Proteins; Biochemical; Biological; Biological Assay; C-terminal; Catalysis; Cells; Cellular biology; Charge; Chromatin; Chromosomes; Clinical; Complex; DNA; DNA Binding; DNA Binding Domain; DNA Integration; Differentiation Inhibitor; Drug resistance; EP300 gene; Elements; Enzymes; Evolution; Face; Foundations; Frequencies; Funding; Future; Genes; Grant; HIV; HIV-1; HIV-1 integrase; HRP-2 protein; Half-Life; Human Chromosomes; In Vitro; Individual; Infection; Integrase; Integrase Inhibitors; Integration Host Factors; Lentivirus Vector; Molecular; Mutation; N-terminal; Nucleoproteins; Null Lymphocytes; PWWP Domain; Paper; Patients; Pharmaceutical Preparations; Play; Positioning Attribute; Proteins; Proteomics; Relative (related person); Research; Reverse Transcription; Role; Site; Specificity; Staging; Structure; System; Techniques; Testing; Transcript; Viral; Viral Physiology; Virus; Virus Assembly; Virus Diseases; Virus Integration; Virus Replication; Work; Zinc Fingers; base; cellular transduction; clinical application; dimer; gene therapy; hepatoma-derived growth factor; improved; in vivo; insight; lens epithelium-derived growth factor; lentiviral-mediated; meetings; monomer; mutant; novel; novel strategies; outcome forecast; preference; public health relevance; research study; resistant strain; structural biology; success; tool; transcriptional coactivator p75; viral DNA

DESCRIPTION (provided by applicant): Millions of people are infected with the AIDS virus HIV-1, and highly active antiviral therapy has made great strides over the past decade to slow virus spread and improve infected individuals' prognoses. Because many of the utilized anti-viral compounds elicit significant side effects, and rapid virus evolution leads to significant numbers of drug resistant strains, there is an ongoing need to discover new drugs to inhibit HIV-1 replication. Such research is driven through detailed understanding of the cellular and molecular biological steps that the virus undergoes to grow. The pivotal step in the viral replication cycle is the integration of the viral DNA made by reverse transcription into a cell chromosome. Integration is catalyzed by integrase, and the first integrase inhibitor was approved for clinical use in 2007. Herein, we will ascertain critical unknown aspects of integrase structure and function. Our work and others have previously highlighted that integrase works in close association with cellular factors to accomplish virus integration. The key factor, lens epithelium-derived growth factor (LEDGF), tethers HIV-1 to active genes during integration, but the underlying mechanistic basis of LEDGF tethering is unknown. This will be deciphered using a variety of cell biology, biochemical, and virological experimental techniques. Even in the complete absence of LEDGF, HIV-1 still favors active genes for integration over random, leading to the hypothesis that other virus-interacting proteins help to guide HIV-1 as it seeks chromosomal sites. The roles of numerous other viral binding proteins in integration will therefore be deciphered. Though it is believed that a tetramer of the integrase protein is the multimer that catalyzes integration, a dearth of detailed structural biology information has limited our overview of its organization. Biochemical and virological experiments will be conducted to define the functional organization of the active integrase tetramer. As LEDGF is the natural tether that guides the virus to sites of integration, novel DNA binding domains will be tested in concert with LEDGF to ascertain the extent that HIV-1 can be directed to new sites for integration. The successful completion of these experiments will uncover fundamental aspects of HIV- 1 integrase structure and function, which will significantly help in the discovery of novel integrase inhibitors. They moreover may open up new ways in which to develop lentiviral vectors for future treatment of patients with genetic therapy. PUBLIC HEALTH RELEVANCE: HIV-1 replication critically relies on integrase activity, the viral enzyme that integrates the reverse transcript into a cell chromosome. This application will uncover novel aspects of integrase structure and function, which will significantly impact ongoing discovery efforts to target and block the pivotal integration step in the virus replication cycle.

描述(申请人提供)：数百万人感染了艾滋病病毒HIV-1，在过去的十年里，高度有效的抗病毒治疗取得了长足的进步，减缓了病毒的传播，改善了感染者的预后。由于许多已使用的抗病毒化合物会产生显著的副作用，而且病毒的快速进化导致了大量的耐药株，因此不断有必要发现抑制HIV-1复制的新药。这种研究是通过详细了解病毒生长所经历的细胞和分子生物学步骤来推动的。病毒复制周期中的关键步骤是将反转录产生的病毒DNA整合到细胞染色体中。整合是由整合酶催化的，2007年第一个整合酶抑制剂被批准用于临床。在这里，我们将确定整合酶结构和功能的关键未知方面。我们的工作和其他人之前已经强调了整合酶与细胞因子密切相关，以完成病毒整合。晶状体上皮源性生长因子(LEDGF)在整合过程中将HIV-1与活性基因捆绑在一起，但LEDGF捆绑的潜在机制尚不清楚。这将使用各种细胞生物学、生化和病毒学实验技术来破译。即使在完全没有LEDGF的情况下，HIV-1仍然倾向于整合活跃的基因，而不是随机整合，这导致了一种假设，即在寻找染色体位置时，其他与病毒相互作用的蛋白质帮助引导HIV-1。因此，许多其他病毒结合蛋白在整合中的作用将被破译。尽管人们认为整合酶蛋白的四聚体是催化整合的多聚体，但缺乏详细的结构生物学信息限制了我们对其组织结构的概述。将进行生化和病毒学实验，以确定活性整合酶四聚体的功能组织。由于LEDGF是将病毒引导到整合部位的天然系绳，因此将与LEDGF一起测试新的DNA结合域，以确定HIV-1可以被引导到新的整合部位的程度。这些实验的成功完成将揭示HIV-1整合酶结构和功能的基本方面，这将大大有助于发现新型整合酶抑制剂。此外，它们还可能开辟开发慢病毒载体的新方法，用于未来对患者进行基因疗法的治疗。 公共卫生相关性：HIV-1复制严重依赖整合酶活性，整合酶是一种将反向转录整合到细胞染色体中的病毒酶。这项应用将揭示整合酶结构和功能的新方面，这将对正在进行的针对和阻止病毒复制周期中关键整合步骤的发现工作产生重大影响。