Assembly of HIV intasomes

项目摘要： 4-结构域原型泡沫病毒（PFV）整合酶（IN）（1，2）和非常小的泡沫病毒（PFV）整合酶（IN）（1，2）的X射线结构。 最近，与病毒DNA和/或病毒靶复合的3-结构域劳斯肉瘤病毒（RSV）IN（3 已确定DNA底物。然而，医学上重要的3- 结构域HIV IN本身或与DNA的复合物仍然难以捉摸。我们建议提供资金， 使用单体野生型（wt）HIV IN或修饰的IN构建体来实现这一目的的方法 困难但意义重大的项目。实现这一重要目标的第一步是 收获HIV IN在高蛋白浓度下组装这些IN-DNA复合物的能力 在~100至150 µM范围内，用于晶体筛选和其他溶液中结构研究。我们 最近成功定义了稳定3-结构域IN-DNA复合物的新组装途径， 重组HIV IN的新的增强溶解度方法的使用将推进我们的研究 问题研究

Project Abstract: The X-ray structures of the 4-domain prototype foamy virus (PFV) integrase (IN)(1,2) and very recently the 3-domain Rous sarcoma virus (RSV) IN (3) in complex with viral DNA and/or viral-target DNA substrates have been determined. Yet, the crystal structure of the medically important 3- domain HIV IN by itself or in complex with DNA is still elusive. We propose to provide the means and methods using monomeric wild type (wt) HIV IN or modified IN constructs to accomplish this very difficult but highly significant project. The first necessary step to achieve this important goal is harvesting the ability of HIV IN to assemble these IN-DNA complexes at high protein concentrations in the ~100 to 150 µM range for crystal screening and other in-solution structural investigations. Our recent success in defining new assembly pathways for stabilizing 3-domain IN-DNA complexes and the use of newly enhanced solubility methods for recombinant HIV IN will advance our investigative studies.