Structural Biology of Retroviral DNA Integration

为了实现生产性感染，HIV必须将其基因组的DNA拷贝插入人类细胞的染色体中。这个复杂的过程是由整合酶，一种由病毒携带的酶协调的。一旦整合完成，病毒基因组就成为细胞染色体中的永久居民。从那里，它将开始产生新的传染性颗粒，或者它可能会保持休眠状态，长时间未被发现。这种融合是逆转录病毒感染持续存在的部分原因。然而，艾滋病毒对融合的依赖也是一个可以利用的弱点。一类新型药物，破坏整合酶的酶活性，称为链转移抑制剂，利用这一弱点来对抗艾滋病毒感染。HIV整合酶的三维原子结构尚不清楚，更不了解的是其在整合过程中的活性形式的结构。目前，缺乏结构信息是链转移抑制剂开发的主要障碍。本计画旨在阐明整合酶的三维结构。我们将分别确定这种蛋白质的原子结构和活性DNA结合形式。为了实现我们的目标，我们将使用X射线晶体学，它允许蛋白质分子的可视化，尽管需要大量的准备工作。特别是，为了确定高分辨率的结构，我们必须获得整合酶与辅助蛋白和/或DNA复合的晶体。我们的研究结果将发表在开放获取期刊上，科学界可以通过公共数据库访问这些数据。我们的研究将产生数据，这将对学术和私人团体的药物发现具有巨大价值，并将有助于降低成本和提高最终治疗的可用性。

To achieve productive infection, HIV must insert a DNA copy of its genome into a chromosome of a human cell. This complex process is orchestrated by integrase, an enzyme carried by the virus. Once integration is complete, the viral genome becomes a permanent resident in a cellular chromosome. From there it will initiate production of new infectious particles or it might stay dormant and undetected for a long period of time. The integration is partly responsible for the notable persistence of retroviral infections. Yet, the dependence of HIV on integration is also an exploitable weakness. A new class of drugs, disrupting enzymatic activity of integrase, called strand transfer inhibitors, takes advantage of this weakness to fight HIV infection. The three-dimensional atomic structure of HIV integrase is not known and even less understood is the architecture of its active form during integration process. Currently, the lack of structural information is the major impediment to the development of strand transfer inhibitors. This project aims to elucidate the three-dimensional structure of integrase. We will determine atomic structures of this protein separately and in active, DNA-bound, form. To achieve our goals we will use X-ray crystallography, which allows visualization of protein molecules, although requiring a significant amount of preparatory work. In particular, to determine high-resolution structures, we will have to obtain crystals of integrase in complex with accessory proteins and/or DNA. Our results will be published in open access journals and the data will be accessible to the scientific community via public databases. Our research will generate dat, which will be of great value for drug discovery by both academic and private groups, and will serve to reduce the costs and improve availability of the eventual treatments.