Elucidating Molecular Mechanisms of Drug Resistance in HIV-1 Protease

阐明 HIV-1 蛋白酶耐药性的分子机制

• 批准号: 8643268
• 负责人: GAIL E FANUCCI
• 金额: $ 26.71万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-04-01 至 2017-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8643268
• 关键词: Accounting; Acquired Immunodeficiency Syndrome; Active Sites; Antiviral Therapy; Binding; Country; Crystallization; Distal; Drug Combinations; Drug resistance; Effectiveness; Electron Spin Resonance Spectroscopy; Electrons; Enzyme Kinetics; Enzymes; Epidemic; Evolution; Generations; Genetic Polymorphism; Goals; HIV; HIV Infections; HIV-1; Impairment; Infection; Kinetics; Life; Literature; Measurement; Measures; Methods; Modeling; Molecular; Molecular Conformation; Mutation; Nelfinavir; Pathway interactions; Patients; Pattern; Peptide Hydrolases; Pharmaceutical Preparations; Pharmacotherapy; Physiologic pulse; Play; Point Mutation; Population; Positioning Attribute; Predisposition; Principal Investigator; Process; Protease Inhibitor; Proteins; Relative (related person); Relaxation; Resistance; Roentgen Rays; Role; Sampling; Site; Spectrum Analysis; Spin Labels; Structure; Surgical Flaps; Testing; Variant; Vertebral column; Viral; Work; base; design; enzyme activity; fitness; flexibility; inhibitor/antagonist; innovation; insight; molecular dynamics; novel; pressure; prevent; programs; public health relevance; response

DESCRIPTION (provided by applicant): HIV-1 infection is a world-wide epidemic. Although current drug therapies are effective at extending patient life, infections continue to spread, especially in non-developed countries, and the emergence of drug resistance has limited the effectiveness of many protease inhibitors. The mechanism by which the evolution of accessory mutations in HIV-1 protease (HIV-1PR) act to recover enzymatic function while imparting cross resistance to numerous inhibitors is the focus of this proposal. We are proposing a novel mechanism, which evokes protein conformational sampling, as a molecular basis to explain how secondary mutations accomplish this task. Specifically we hypothesize that mutations combine in patterns that alter HIV-1PR conformational sampling among four nominal states; namely, the closed, the semi- open, curled/tucked and wide-open such that (1) enzyme function is recovered by the secondary mutations when they combine to stabilize the semi-open conformation to a percentage observed in a native enzyme (e.g. inhibitor naive sequence) and (2) that cross resistance emerges when the mutations combine to stabilize "open-like" states such as the wide-open and curled/tucked conformations while concomitantly destabilizing the closed state population. It is through an innovative application of pulsed electron paramagnetic spectroscopy, which our lab has pioneered over the last five years that we measure conformational sampling ensembles in HIV-1PR. Within the aims of this proposal, we will make correlations among changes in HIV-1PR conformational sampling ensembles to enzymatic parameters, inhibition constants, inhibitor susceptibility, and viral fitness for numerous HIV-1PR variants. From these results, if our hypothesis is correct, we will provide a molecular level understanding of how secondary mutations elicit their effects on HIV-1PR. Additionally, these studies will also provide insights into explaining why the patterns of secondary mutation evolution against protease inhibitors are divergent in non-B subtypes. Finally, we will target numerous variants that show cross-resistance and increased conformational sampling of the open-like states for structure determination. Structures with more open-like conformations can serve as targets for the rational design of the new inhibitors for treatment of extremely resistant HIV-1PR variants.

描述（由申请人提供）：HIV-1感染是一种世界性流行病。尽管目前的药物治疗在延长患者生命方面是有效的，但感染继续蔓延，特别是在非发达国家，并且耐药性的出现限制了许多蛋白酶抑制剂的有效性。HIV-1蛋白酶（HIV-1 PR）中辅助突变的进化作用恢复酶功能，同时赋予对许多抑制剂的交叉耐药性的机制是本提案的重点。我们提出了一种新的机制，它唤起蛋白质构象采样，作为分子基础来解释二级突变如何完成这一任务。具体地说，我们假设突变联合收割机的模式，改变HIV-1 PR的构象抽样之间的四个名义状态;即封闭式，半开放式，卷曲/折叠和完全开放，使得（1）当二级突变联合收割机组合以将半开放构象稳定到在天然酶中观察到的百分比时，酶功能通过二级突变恢复（例如抑制剂初始序列）和（2）当突变联合收割机组合以稳定“开放样”状态（例如大开放和卷曲/折叠构象）同时伴随使封闭状态群体不稳定时出现交叉抗性。正是通过脉冲电子顺磁光谱的创新应用，我们的实验室在过去五年中开创了我们测量HIV-1 PR中构象采样集合的先河。在本提案的目标，我们将在HIV-1 PR构象采样合奏酶参数，抑制常数，抑制剂的敏感性，和病毒的健身众多的HIV-1 PR变异体之间的相关性。从这些结果中，如果我们的假设是正确的，我们将提供一个分子水平的理解如何继发突变引起他们对HIV-1 PR的影响。 此外，这些研究还将提供见解，解释为什么蛋白酶抑制剂的二次突变进化模式在非B亚型中是不同的。最后，我们将针对许多变体，这些变体显示出交叉抗性，并增加了开放样状态的构象采样以进行结构测定。具有更多开放样构象的结构可以作为合理设计用于治疗极端耐药的新抑制剂的靶点。 HIV-1 PR变异体。