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: 8073053
• 负责人: Jamil Subhi Saad
• 金额: $ 36.26万
• 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 2015-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8073053
• 关键词: Affect; Affinity; Antiviral Agents; Binding; Biochemical; Biological Assay; Calcium; Calmodulin; Cell membrane; Cell physiology; Cells; Complex; Cytokine Inducible SH2-Containing Protein; Cytoplasm; Data; Development; Drug Design; Electrostatics; Gagging; HIV; HIV Infections; HIV-1; Histidine; Human; Infection; Integration Host Factors; Knowledge; Lead; Lecithin; Life; Lipids; Location; Membrane; Membrane Lipids; Methods; Micelles; Modeling; Molecular; Mutagenesis; Nuclear Magnetic Resonance; Nucleocapsid; Pathway interactions; Peptides; Phase; Phosphatidylethanolamine; Phosphatidylinositol 4,5-Diphosphate; Phosphatidylserines; Phospholipids; Property; Protein Binding; Proteins; Public Health; Research; Resolution; Role; Safety; Signal Pathway; Signal Transduction; Specificity; Staging; Structure; Techniques; Therapeutic Agents; Virus Replication; base; design; gag Gene Products; in vivo; membrane assembly; membrane model; novel strategies; particle; pathogen; phosphatidylethanolamine; protein complex; public health relevance; structural biology; three dimensional structure; tool; trafficking

DESCRIPTION (provided by applicant): Within cells proteins are targeted to well-defined locations by specific interactions with cellular constituents such as proteins and phospholipids. During the late phase of HIV-1 infection, Gag polyproteins are transported to the plasma membrane (PM) for assembly. Using nuclear magnetic resonance (NMR) we have previously defined the molecular basis of one component of Gag targeting to the PM, the interaction of the matrix (MA) domain with phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2). Our preliminary data indicate that Gag also forms biologically important interactions with cellular lipids and proteins implicated in efficient particle assembly. We show that MA interacts directly with phosphatidylserine (PtdSer), phosphatidylcholine (PtdCho) and phosphatidylethanolamine (PtdEtn), suggesting that Gag targeting and assembly on the PM proceed via a dual engagement of the MA domain with PI(4,5)P2 and major membrane lipids. A number of cellular proteins have been proposed to facilitate Gag intracellular trafficking and targeting to the PM. HIV-1 Gag was shown to interact and co-localize with calmodulin (CaM) in the cytoplasm. In addition, more recent studies have identified the suppressor of cytokine signaling 1 (SOCS1) as an inducible host factor during HIV infection, which regulates the late stages of the virus replication pathway through direct interactions with the Gag protein. However, the molecular mechanism by which Gag interacts with these factors has yet to be established. Preliminary studies demonstrate that HIV-1 MA interacts directly with CaM in a calcium- dependent manner, suggesting that HIV-1 hijacks CaM cell-signaling pathway to facilitate Gag trafficking. In addition, we have obtained evidence for direct interactions between SOCS1 and the MA protein. In this proposal, we will employ biochemical, biophysical and structural biology tools to identify key protein-protein and protein-lipid interactions involved in the molecular mechanism governing HIV-1 Gag intracellular trafficking and assembly. Our three main aims are: (i) to determine the precise molecular mechanism for Gag binding to the PM. We will determine at the structural level how various lipid constituents interact with the MA protein and will identify the functional importance of the hydrophobic insertion, specific and non- specific electrostatic interactions, (ii) to identify the functional role of CaM in Gag trafficking and assembly, and (iii) to elucidate the structural requirements for SOCS1-Gag interactions. The results generated by this research will offer a better understanding of how HIV-1 Gag interacts with cellular constituents at the atomic level, which could lead to new approaches to rational design of new antiviral therapeutic agents that inhibit Gag trafficking and assembly. PUBLIC HEALTH RELEVANCE: HIV remains the most deadly pathogen that threatens human's life and the lack of efficient cure poses a major problem in public health and safety. Major efforts are now being focused on understanding how HIV interacts with the host cell during the replication cycle. The proposed studies will help in understanding the precise molecular mechanism of HIV Gag trafficking and assembly, which may aid in the development of new strategies for antiviral drug design.

描述（由申请人提供）：通过与细胞成分（例如蛋白质和磷脂）的特定相互作用，细胞蛋白质蛋白质内的蛋白质在定义明确的位置。在HIV-1感染的后期，将GAG多蛋白转移到质膜（PM）中进行组装。使用核磁共振（NMR），我们先前已经定义了靶向PM的一个成分的分子基础，即矩阵（MA）结构域与磷脂酰肌醇-4,5-双磷酸（PI（4,5）P2）的相互作用。我们的初步数据表明，GAG还与涉及有效颗粒组装的细胞脂质和蛋白质形成了重要的生物学相互作用。我们表明，MA与磷脂酰甲酯（PTDSER），磷脂酰胆碱（PTDCHO）和磷脂酰乙醇胺（PTDETN）直接相互作用，这表明PM上的PM靶向和组装通过MA Domain与PI（4,5）P2和Major Membrane Lipids进行双重订婚。已经提出了许多细胞蛋白，以促进细胞内运输并靶向PM。 HIV-1插入被证明与细胞质中的钙调蛋白（CAM）相互作用并共定位。此外，最近的研究将细胞因子信号传导1（SOCS1）的抑制剂抑制为HIV感染期间的诱导宿主因子，该因素通过与GAG蛋白的直接相互作用来调节病毒复制途径的晚期。然而，GAG与这些因素相互作用的分子机制尚未确定。初步研究表明，HIV-1 MA以钙依赖性方式直接与CAM相互作用，这表明HIV-1劫持了CAM CAM细胞信号途径以促进GAG运输。此外，我们还获得了SOCS1与MA蛋白之间直接相互作用的证据。在此提案中，我们将采用生化，生物物理和结构生物学工具来鉴定有关控制HIV-1 GAG GAG细胞内运输和组装的分子机制中涉及的关键蛋白质蛋白质和蛋白质脂质相互作用。我们的三个主要目的是：（i）确定与PM结合的GAG结合的精确分子机制。我们将在结构级别确定各种脂质成分如何与MA蛋白相互作用，并确定疏水插入，特异性和非特异性静电相互作用的功能重要性，（ii）确定CAM在GAG运输和组装中的功能作用，以及（III）以阐明SOCS1-GAG相互作用的结构需求。这项研究产生的结果将更好地了解HIV-1 GAG如何与原子水平上的细胞成分相互作用，这可能会导致新的抗病毒治疗剂理性设计的新方法，从而抑制GAG贩运和组装。 公共卫生相关性：艾滋病毒仍然是威胁人类生命和缺乏有效治愈的最致命的病原体，这在公共卫生和安全方面构成了主要问题。现在，主要的努力集中在理解在复制周期中艾滋病毒如何与宿主细胞相互作用。拟议的研究将有助于理解艾滋病毒插科打贩运和组装的精确分子机制，这可能有助于制定新的抗病毒药药设计策略。