Integrase Structural Virology

 描述(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，而人类免疫缺陷病毒1型(HIV-1)的整合酶是用于治疗艾滋病患者的高效抗逆转录病毒疗法的共同靶点。整合酶链转移抑制剂(INSTI)自2007年以来一直在临床使用，这一更新应用的先前迭代关键地发现了它们的作用机制。INSTI的靶点是整合酶-病毒DNA核蛋白复合体，也称为内切酶，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HIV-1内切酶的高分辨结构尚未被报道，但我们已经报道了大量的原型泡沫病毒(PFV)内切酶的结构，它也被INSTI抑制。利用这个模型，我们先前描述了INSTI的工作原理是将病毒DNA的关键脱氧腺苷残基及其相关的3‘-羟基亲核体从整合酶活性部位排出，解除整合机制的武装。在当前的资助期间，我们延长了这一观察范围，发现INSTI是对DNA链转移反应的攻击和离开基团的化学结构的模拟。在这里，我们提供了一种新的逆转录病毒内切酶结构的初步数据，我们将使用这个新的结构信息与PFV内切酶的结构一起来完善我们的HIV-1内切酶的工作模型，这是临床上相关的INSTI靶标。我们发现，自INSTI以来最耐人寻味的整合酶药物类别，变构整合酶抑制剂(ALLINIS)，抑制HIV-1颗粒成熟，暗示整合酶在HIV-1颗粒形态发生中的结构作用。整合酶在形成具有感染性的HIV-1结构中的作用将使用各种生化和基因技术来阐明。这一系列研究的最终结果将是HIV-1连接体的生物现实模型，以帮助新的INSTI开发，以及对Allini作用机制的敏锐愿景，这将为这一新的和重要的抗HIV药物类别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.