OrthoSwith Probes for High Throughput Screening

描述（由申请人提供）：该项目旨在创建和使用工具来筛选大型化合物库，以发现和评估HIV-1的三个靶点的候选药物。它们是(1)rnanuclecapsid （NC）蛋白复合物，(2)TAR RNA-Tat蛋白- cyclint1蛋白复合物，以及(3)Vif蛋白。目前的艾滋病治疗中还没有这些靶点的抑制剂，在这个项目的第一阶段，我们成功地创造了一种分子开关，它在NC存在的情况下改变其状态，并在一种模仿病毒RNA包装信号的抑制剂存在的情况下开关回来。该抑制剂在几秒钟内引起荧光增加，这在候选药物的高通量筛选（HTS）中很容易监测。这是很容易和明确的验证，以1:1的比例发生的蛋白质和开关组分（或不），并确定亲和常数，K/d。我们提出了有限的原型，并创建HTS-和kdkit，其他人可以使用它来发现和改进候选药物，以中断HIV-I中的NC， Tat和Vif功能。在未来，类似的试验将会用新药靶向所有15种HIV蛋白。

DESCRIPTION (provided by applicant): This project aims at creating and using the tools to screen large libraries of chemical compounds to find and evaluate drug candidates for three targets in HIV-1. These are (1) the RNAnucleocapsid (NC) protein complex, (2) the TAR RNA-Tat protein-CyclinT1 protein complex, and (3) the Vif protein. No inhibitors of these targets are available in current AIDS therapies, in phase I of this project we succeeded in creating a molecular switch that changes its state in the presence of NC, and switches back in the presence of an inhibitor that mimics the viral RNA packaging signal. The inhibitor causes an increase in fluorescence within seconds that is easily monitored in high throughput screens (HTS) of candidate drugs. It is easy and unambiguous to verify that the complex occurs with a 1:1 ratio of protein and switch components (or not), and to determine the affinity constant, K/d. We propose limited prototyping and to create HTS- and Kdkits that others can use to discover and refine drug candidates to interrupt NC, Tat, and Vif function in HIV-I. In the future, similar assays will target all fifteen HIV proteins with new drugs.