STRUCTURE/FUNCTION OF INTEGRASE

INTEGRASE的结构/功能

• 批准号: 6373561
• 负责人: ANNA MARIE SKALKA
• 金额: $ 44.64万
• 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 2002-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6373561
• 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），催化新近的插入 将病毒DNA合成到宿主细胞染色体中。 这个事件是 正常病毒复制所必需的。 因此，像HIV-1相反 转录酶和蛋白酶，IN是抑制的潜在目标 预防或治疗艾滋病。 描述的研究的主要目标 在此建议中是了解HIV-1的分子结构， 单独且与其DNA底物复杂化，因此 抑制剂可能以理性的基础为基础。 三个具体目标是 提议：第一个特定目的描述了不同的结构分析 在有金属辅因子的情况下，域和全长 使用X-模型底物抑制剂和单克隆抗体片段 射线晶体学和NMR。 第二个特定目标将测试特定 结构数据建议的组织和功能模型， 使用位置定向的诱变和In的化学修饰。 这 第三个特定目的描述了新颖的突触末端病毒DNA的使用 底物研究功能中如何与DNA相互作用 催化协调加工反应的复合物。 特定的 预测DNA弯曲和部分放松的模型将被测试， 使用DNA足迹技术。 在此描述的实验 提案采用广泛的最先进的分子遗传， 生化和生物物理方法以及新型底物和 免疫试剂。 他们还以丰富的经验为基础 与人类（艾滋病毒）和鸟类（ASV）的PI和合作者的合作者 病毒整合系统，以及将ASV用作有价值的能力 比较基础。 获得的知识应有助于描述 特定于艾滋病毒的功能。 这些研究的结果提供了 抑制HIV整合和发育的新分子靶标 艾滋病疗法。