STRUCTURE/FUNCTION OF INTEGRASE

INTEGRASE的结构/功能

• 批准号: 6170297
• 负责人: ANNA MARIE SKALKA
• 金额: $ 43.34万
• 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 2002-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6170297
• 关键词: DNA binding protein; DNA footprinting; X ray crystallography; antiAIDS agent; avian sarcoma virus; chemical models; drug design /synthesis /production; enzyme activity; human immunodeficiency virus 1; integrase; laboratory mouse; laboratory rabbit; laboratory rat; nuclear magnetic resonance spectroscopy; protein structure function; site directed mutagenesis

The retroviral enzyme, integrase (IN), catalyzes the insertion of newly synthesized viral DNA into the host cell chromosome. This event is required for normal viral replication. Thus, like HIV-1 reverse transcriptase and protease, IN is a potential target for inhibition in prevention or treatment of AIDS. The major goal of research described in this proposal is to understand the molecular structure of HIV-1 IN, alone and in complex with its DNA substrates, so that design of inhibitors may build upon a rational foundation. Three Specific Aims are proposed: The first Specific Aim describes structural analyses of distinct domains and full length IN, in the presence of metal cofactors, selected model substrate inhibitors and monoclonal antibody fragments, using X- ray crystallography and NMR. The second Specific Aim will test specific models of IN organization and function suggested by the structural data, using site-directed mutagenesis and chemical modification of IN. The third Specific Aim describes the use of a novel, synapsed-end viral DNA substrate to investigate how IN interacts with DNA in a functional complex that catalyzes coordinated processing reactions. A specific model that predicts DNA bending and partial unwinding will be tested, using DNA footprinting techniques. Experiments described in this proposal employ a broad range of state-of-the-art molecular genetic, biochemical, and biophysical methods, as well as novel substrates and immunological reagents. They also build upon the extensive experience of the PI and collaborators with both the human (HIV) and avian (ASV) viral integration systems, and the ability to utilize ASV IN as a valuable base of comparison. The knowledge gained should help to delineate features that are specific to HIV. The results of these studies provide new molecular targets for inhibition of HIV integration and development of AIDS therapies.

逆转录病毒酶，整合酶（IN），催化新的插入 将病毒DNA合成到宿主细胞染色体中。 本次活动是 正常病毒复制所需。 因此，就像 HIV-1 逆转一样 转录酶和蛋白酶，IN 是抑制的潜在靶点 预防或治疗艾滋病。 描述的研究的主要目标 这个提案的目的是了解HIV-1 IN的分子结构， 单独或与其DNA底物复合，因此设计 抑制剂可以建立在合理的基础上。 三个具体目标是 建议：第一个具体目标描述了不同的结构分析 结构域和全长IN，在金属辅因子存在下，选择 模型底物抑制剂和单克隆抗体片段，使用 X- 射线晶体学和核磁共振。 第二个具体目标将测试具体 结构数据建议的 IN 组织和功能模型， 使用IN的定点诱变和化学修饰。 这 第三个具体目标描述了新型突触端病毒 DNA 的使用 底物以研究 IN 如何与 DNA 相互作用 催化协调加工反应的复合物。 具体的 将测试预测 DNA 弯曲和部分解旋的模型， 使用 DNA 足迹技术。 本文描述的实验 建议采用广泛的最先进的分子遗传学， 生物化学和生物物理方法，以及新的底物和 免疫试剂。 他们还以丰富的经验为基础 PI 和合作者与人类 (HIV) 和禽类 (ASV) 病毒整合系统，以及利用 ASV IN 作为有价值的 比较的基础。 所获得的知识应该有助于描述 HIV 特有的特征。 这些研究的结果提供了 抑制艾滋病毒整合和发展的新分子靶点 艾滋病疗法。