Integrase Structural Virology

 描述（由申请人提供）：由病毒整合酶蛋白催化的整合是所有逆转录病毒生命周期中的重要步骤，并且人类免疫缺陷病毒1型（HIV-1）的整合酶是用于治疗AIDS患者的高效抗逆转录病毒疗法的常见靶标。整合酶链转移抑制剂（INSTIs）自2007年以来一直在临床使用，此次更新申请的前期迭代批判性地发现了其作用机制。INSTI的靶点是整合酶-病毒DNA核蛋白复合物，也称为整合体，通过研究药物靶点的详细三维结构，极大地促进了对治疗作用机制的理解。虽然高分辨率的结构的HIV-1 intasome还没有被报道，我们已经报道了许多结构的原型泡沫病毒（PFV）intasome，这也是抑制的INSTI。使用该模型，我们先前描述了INSTI通过从整合酶活性位点喷射病毒DNA的关键脱氧腺苷酸残基及其相关的3 '-羟基亲核试剂，解除整合机制而起作用。在目前的资助期间，我们扩展了这一观察，发现INSTI是DNA链转移反应的化学攻击和离去基团的结构模拟物。在此，我们提供了一种新的逆转录病毒intasome结构的初步数据，我们将使用这种新的结构信息与PFV intasome的结构一起完善我们的HIV-1 intasome工作模型，临床相关的HIV-1靶点。我们已经发现，自INSTI以来最有趣的整合酶药物类别，变构整合酶抑制剂（ALLINIS），抑制HIV-1颗粒成熟，这意味着整合酶在HIV-1颗粒形态发生中的结构作用。整合酶在形成感染性HIV-1结构中的作用将使用各种生物化学和遗传学技术来阐明。这一系列的研究将最终为HIV-1整合体提供一个生物学上现实的模型，以帮助开发新的ALLINI，并对ALLINI的作用机制有一个敏锐的认识，这将为这种新的重要的抗HIV药物的临床开发提供信息。

 DESCRIPTION (provided by applicant): Integration, catalyzed by the viral integrase protein, is an essential step in the life cycle of all retroviruses, and the integrase enzyme of human immunodeficiency virus type 1 (HIV-1) is a common target of the highly active antiretroviral therapies that are used to treat AIDS patients. Integrase strand transfer inhibitors (INSTIs) have been in clinical use since 2007, and the prior iteration of this renewal application critically discovered their mechanism of action. The target of the INSTIs is the integrase-viral DNA nucleoprotein complex, also known as the intasome, and the understanding of the mechanism of therapeutic action is greatly facilitated through the study of detailed 3- dimensional structure of drug targets. Although a high-resolution structure of the HIV-1 intasome has yet to be reported, we have reported numerous structures for the prototype foamy virus (PFV) intasome, which is also inhibited by the INSTIs. Using this model, we previously described that INSTIs work by ejecting the critical deoxyadenylate residue of viral DNA and its associated 3'-hydroxyl nucleophile from the integrase active site, disarming the integration machinery. During the current funding period we extended this observation to discover that INSTIs are structural mimics of the chemical attacking and leaving groups of the DNA strand transfer reaction. Herein we provide preliminary data for a new retroviral intasome structure, and we will use this new structural information together with the structure of the PFV intasome to refine our working model of the HIV-1 intasome, the clinically relevant INSTI target. We have discovered that the most intriguing integrase drug class since the INSTIs, the allosteric integrase inhibitors (ALLINIS), inhibits HIV-1 particle maturation, implying a structural role for integrase in HIV-1 particle morphogenesis. The role of integrase in forming the infectious HIV-1 structure will be elucidated using a variety of biochemical and genetic techniques. This line of research will culminate with a biologically realistic model for the HIV-1 intasome to aid novel INSTI development and with an acute vision of the mechanism of ALLINI action, which will inform the clinical development of this new and important anti-HIV drug class.