Structural Basis for HIV-1 Gag Interactions with Cellular Constituents

HIV-1 Gag 与细胞成分相互作用的结构基础

• 批准号: 9304625
• 负责人: Jamil Subhi Saad
• 金额: $ 15万
• 依托单位: UNIVERSITY OF ALABAMA AT BIRMINGHAM
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-05-15 至 2019-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9304625
• 关键词: Acquired Immunodeficiency Syndrome; Address; Affinity; Antiviral Agents; Avian Sarcoma Viruses; Binding; Binding Proteins; Biochemical; Capsid; Cell membrane; Cholesterol; Cytoplasm; Cytoplasmic Tail; Data; Development; Drug Design; Elements; Engineering; Genetic; HIV; HIV-1; Human; Hydrophobicity; Infection; Knowledge; Lead; Life; Light; Masks; Mediating; Membrane; Membrane Lipids; Membrane Proteins; Molecular; Molecular Conformation; Molecular Models; N-terminal; Nucleocapsid; Outcome; Penetration; Phase; Process; Production; Proteins; Public Health; RNA; Recruitment Activity; Retroviridae; Role; Safety; Site; Structural Genes; Structure; Suggestion; Surface Tension; Techniques; Therapeutic Agents; Virion; Virus Assembly; base; biophysical techniques; combat; env Gene Products; gag Gene Products; in vivo; innovation; insight; mimetics; molecular modeling; novel; novel strategies; novel therapeutic intervention; particle; pathogen; protein complex; reconstitution; tool

During the late phase of HIV-1 infection, the Gag protein is synthesized and folds in the cytoplasm into an assembly inactive form in which not only the nucleocapsid (NC) domain but also the matrix (MA) domain interacts with RNA. As a consequence of this "folded" conformation, the capsid (CA) domain becomes incapable of assembling. During or upon transport to the plasma membrane (PM) site of assembly, a conformational switch occurs in which the interaction of MA with NC is exchanged for an interaction of MA with PM components. This switch liberates the CA domains for assembly and this conformational rearrangement, essential for activation of Gag, is driven by the binding affinity of specific PM components for MA. To understand this essential switch, it is fundamentally important to elucidate at the molecular level the interaction of MA with PM components. Interestingly, the Gag proteins of closely related retroviruses such as avian sarcoma virus (ASV) contain structural domains with functions essentially identical to those of HIV-1 Gag; however, the mechanisms for Gag assembly on the PM appear to be different. Understanding the molecular basis of this switch will not only shed light on the assembly of HIV-1 but is likely to provide insight into the control of assembly in other retroviruses that assemble at the PM. Subsequent to assembly on the PM, the Gag proteins recruit the envelope (Env) protein for incorporation into virus particles. There is mounting evidence that incorporation of the Env protein in nascent virions is mediated by interactions between the MA domain of Gag and the cytoplasmic tail (CT) of gp41. The mechanism by which Gag mediates Env incorporation is not known. In this proposal, we will employ NMR, biophysical, biochemical and in vivo tools to provide a macromolecular picture of how the Gag protein binds to the PM, and to determine the molecular elements for Gag-gp41 interactions that mediate Env incorporation into virus particles. In Aim 1, we will determine the precise molecular mechanism for HIV-1 Gag binding to the PM. We have devised innovative approaches to characterize interactions between MA and membrane mimetics. In Aim 2, we will identify the molecular elements of ASV Gag assembly on the PM. We present evidence for a novel molecular switch in ASV MA upon binding to PM components. In Aim 3, we will characterize the interactions between HIV-1 MA and gp41 CT. We devised a new strategy to reconstitute and mimic the physiologically relevant conditions for interactions. These studies will provide details on the molecular switches that activate Gag assembly on the PM and help in the recruitment of the Env protein to assembly sites. We hope that the outcome of this proposal will help in the development of new antiviral therapeutic agents that inhibit assembly and virus production.

在HIV-1感染的后期，合成了GAG蛋白并在细胞质中折叠成一个 组装非活动形式不仅在其中核素（NC）结构域，还包括矩阵（MA）域 与RNA相互作用。由于这种“折叠”构象的结果，capsid（ca）域变为 无法组装。在质膜（PM）组件位点运输期间或运输时 发生构象开关，其中MA与NC的相互作用交换以作为MA的相互作用 使用PM组件。此开关解放了CA域的组装和这种构象 重排，对于激活GAG必不可少 对于马。要了解这种基本转换，在分子水平上阐明这一点至关重要 MA与PM组件的相互作用。有趣的是，密切相关逆转录病毒的GAG蛋白 例如禽肉瘤病毒（ASV）包含具有与之相同的功能的结构域 HIV-1插科打;但是，PM上的插科打组件的机制似乎不同。理解 该开关的分子基础不仅会阐明HIV-1的组件，而且可能会提供 深入了解在PM组装的其他逆转录病毒中的组装控制。在组装之后 在PM上，GAG蛋白募集了包膜（ENV）蛋白以掺入病毒颗粒中。有 越来越多的证据表明，将ENV蛋白掺入新生病毒体中是由相互作用介导的 在GP41的GAG和细胞质尾（CT）之间。插科打的机制 中间设想成立尚不清楚。在此提案中，我们将采用NMR，生物物理，生化 和体内工具，以提供大分子图片，说明GAG蛋白如何与PM结合以及与PM结合 确定GAG-GP41相互作用的分子元素，以介导设想融合到病毒中 颗粒。在AIM 1中，我们将确定HIV-1 GAG与PM结合的精确分子机制。 我们设计了创新的方法来表征MA和膜模拟物之间的相互作用。 在AIM 2中，我们将确定PM上ASV GAG组件的分子元件。我们提供证据 与PM成分结合后，ASV MA中的一种新型分子开关。在AIM 3中，我们将表征 HIV-1 MA和GP41 CT之间的相互作用。我们制定了一种重新建立和模仿的新战略 相互作用的生理相关条件。这些研究将提供有关分子的细节 激活PM上插科打插件的开关并有助于募集Env蛋白来组装 站点。我们希望该提案的结果将有助于开发新的抗病毒治疗 抑制组装和病毒产生的药物。