Structure and Function of Integrase

Integrase的结构和功能

• 批准号: 8463943
• 负责人: ANNA MARIE SKALKA
• 金额: $ 41.95万
• 依托单位: RESEARCH INST OF FOX CHASE CAN CTR
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-08-01 至 2017-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8463943
• 关键词: AIDS therapy; AIDS/HIV problem; Active Sites; Address; Affect; Amino Acid Substitution; Architecture; Avian Sarcoma Viruses; Binding; Biological Assay; Catalysis; Chemicals; Clinical; Clinical Trials; Complement; Coupled; DNA; DNA Binding; Data; Data Analyses; Deuterium; Development; Drug Targeting; Enzymes; FDA approved; Fluorescence; Funding; HIV; HIV Integrase; HIV drug resistance; HIV-1; Hydrogen; Integrase; Integrase Inhibitors; Knowledge; Lead; Length; Maps; Mass Spectrum Analysis; Measures; Methods; Modeling; Molecular Conformation; Monitor; Pharmaceutical Preparations; Positioning Attribute; Property; Proteins; Publishing; Reaction; Research; Roentgen Rays; Role; Shapes; Site; Solutions; Spumavirus; Structural Models; Structure; Testing; Therapeutic; Viral; Viral Physiology; Virus; base; crosslink; design; dimer; drug development; in vitro activity; inhibitor/antagonist; insight; monomer; mutant; novel strategies; novel therapeutics; physical model; prototype; research study; screening; single-molecule FRET; small molecule libraries; success; viral DNA; viral resistance

DESCRIPTION (provided by applicant): Integrase (IN) is one of three virus-encoded enzymes that are essential for retroviral replication and a validated target for the development of drugs t treat HIV/AIDS. Although there has been success in developing clinically useful drugs that block the final step in the integration reaction, the so-called strand transfer inhibitors, there is a continuing need to augment or replace existing IN therapeutics as viral resistance is encountered. A detailed knowledge of all aspects of the structure, assembly, and catalysis by HIV-1 IN, will reveal unexploited vulnerabilities and novel strategies for inhibiting this critical enzyme. In the current funding period, we applied small angle X-ray scattering (SAXS) and protein-protein cross-linking methods to obtain the first experimentally-derived models of full-length unliganded apo-IN monomers and dimers in solution, using avian sarcoma virus (ASV) IN. The results revealed a dimer architecture (called a reaching dimer) that was previously unsuspected. The configuration of the reaching dimer resembles that of the viral DNA-binding, "inner" dimer in the crystal structure of the prototype foamy virus (PFV) IN. From these and other data, we have constructed a structural model for an HIV IN reaching dimer, which we hypothesize is pre-positioned to interact with viral DNA ends. In Aim 1 of this competitive renewal we propose to test this model by determining the solution structures of monomers, dimers, and tetramers of HIV IN, using methods successfully employed with ASV IN. We will identify the interactions that stabilize HIV dimers and determine the effects of substrate binding on their conformation. In Aim 2 we will identify compounds that alter the stability of HIV apo-IN dimers and inhibit the conformational changes that are required for IN function. The results of our studies will provide critical new information concerning HIV IN structure and function, and contribute to the design of new, allosterically-acting drugs that can complement the active site inhibitors now in clinical use.

描述（由申请人提供）：综合酶（in）是三种对逆转录病毒复制至关重要的酶之一，也是验证的药物开发的靶标T治疗HIV/AIDS。尽管开发临床上有用的药物已经成功阻止了整合反应的最后一步，即所谓的链转移抑制剂，但由于遇到病毒抗性，持续需要增加或替代治疗剂中存在的疗法。 HIV-1 IN的结构，组装和催化的各个方面的详细知识将揭示未开发的脆弱性和抑制这种关键的新型策略 酶。在当前的资金期间，我们使用鸟类肉瘤病毒（ASV）在溶液中应用了小角度X射线散射（SAXS）和蛋白质 - 蛋白质交联方法，以获取第一个实验衍生的实验衍生的模型，该模型使用鸟类肉瘤病毒（ASV）。结果揭示了以前没有使用的二聚体体系结构（称为到达二聚体）。到达二聚体的构型类似于病毒DNA结合的构型，在原型泡沫病毒（PFV）In的晶体结构中的“内”二聚体。从这些数据和其他数据中，我们在达到二聚体时构建了一个HIV的结构模型，我们假设该模型已预先定位以与病毒DNA末端相互作用。在此竞争性更新的目标1中，我们建议使用ASV IN成功使用的方法来确定HIV IN的单体，二聚体和四聚体的溶液结构来测试该模型。我们将确定稳定HIV二聚体的相互作用，并确定底物结合对其构象的影响。在AIM 2中，我们将确定改变HIV Apo-In二聚体稳定性并抑制功能中所需的构象变化的化合物。我们的研究结果将提供有关结构和功能中HIV的关键新信息，并有助于设计新的，变构作用的药物，这些药物现在可以与现在的临床用途相互补充活跃部位抑制剂。