Function and mechanism of the HCV p7 channel and its therapeutic potential

HCV p7通道的功能、机制及其治疗潜力

• 批准号: 9198039
• 负责人: JAMES Jeiwen CHOU
• 金额: $ 46.4万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-01-01 至 2019-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9198039
• 关键词: Address; Adopted; Antiviral Agents; Architecture; Binding; Binding Sites; Biological Assay; Biology; Calcium; Calcium Channel; Capsid; Cations; Chronic; Complex; Consensus; Data; Drug Interactions; Elements; Environment; Hepatitis C; Integral Membrane Protein; Investigation; Ions; Knowledge; Life Cycle Stages; Link; Liver diseases; Measurement; Mediating; Membrane; Methods; Molecular; Molecular Conformation; Molecular Virology; Mutagenesis; Mutation; Names; Nature; Pathogenicity; Pharmaceutical Chemistry; Pharmaceutical Preparations; Production; Proteins; Protons; RNA; RNA Viruses; Reading; Reporting; Research; Resistance; Role; Site; Structure; System; Therapeutic; Titrations; Viral; Viral Genome; Viral Proteins; Virion; Virus; Virus Assembly; Virus Diseases; Work; base; drug development; experimental study; extracellular; inhibitor/antagonist; insight; interdisciplinary approach; molecular dynamics; mutant; novel; protein protein interaction; public health relevance; small molecule; structural biology; synergism; virology

 DESCRIPTION (provided by applicant): Many viral genomes encode small integral membrane proteins that form channels in membrane, and they conduct protons, cations, and other molecules across the membrane barrier to aid various steps of viral entry and maturation. These viral proteins, collectively named viroporins, are crucial for viral pathogenicity and have been pursued as antiviral targets. The viroporin p7 encoded by the Hepatitis C virus (HCV) genome is one of the more important viral channels that has been validated as a target for developing drugs for treating HCV infections. p7 is an integral membrane protein that oligomerizes to form channels with cation selectivity, for Ca2+ over other cations; it has been shown to facilitate assembly and egress of infectious virions. The name "viroporin" assigned to p7 suggests a simple pore until our lab showed in Ouyang et al Nature 2013 that p7 adopts a rather sophisticated mode of hexameric assembly. The novel architecture implies a new channel mechanism developed by the HCV, but at present, the structural bases for Ca2+ selectivity and ion conduction are unknown. It has been shown that blocking p7 channel activity reduces production of infectious viral progeny, and several compounds have already been identified. These drug interactions should provide useful information for rational drug development, but there is no information on how and where these compounds act in the p7 channel. Furthermore, despite the consensus that p7 channel activity is important for virus production, it remains unclear in which of the steps of assembly and release is the role of channel activity required. We will capitalize on the recent p7 channel structure and utilize a multidisciplinary approach involving structural biology, channel recording, molecular virology, and medicinal chemistry to address these key questions. In Aim 1, we will determine the structure of the p7 channel in bicelles, identify key residues important for cation binding and conduction, and investigate conformational exchange relevant to the conduction mechanism. In Aim 2, we will separate the channel activity of p7 from its role in protein-protein interaction through mutagenesis to understand the role of p7 channel activity during virus infection. Finally in Aim 3, we will identiy the inhibitor binding sites, delineate mechanism of inhibition, and explore strategies to develop new inhibitors. The knowledge to be gained from the proposed research may give rise to new opportunities for developing compounds for treating HCV infections.

 描述(申请人提供)：许多病毒基因组编码小的完整的膜蛋白，在膜上形成通道，它们引导质子、阳离子和其他分子跨过膜屏障，帮助病毒进入和成熟的各个步骤。这些病毒蛋白，统称为病毒孔蛋白，对病毒的致病性至关重要，一直被用作抗病毒靶标。丙型肝炎病毒基因组编码的病毒孔蛋白p7是一种重要的病毒通道，已被证实是开发治疗丙型肝炎病毒感染药物的靶点。P7是一种完整的膜蛋白，它通过寡聚形成具有阳离子选择性的通道，使钙离子高于其他阳离子；它被证明有助于感染性病毒粒子的组装和排出。在我们的实验室在2013年的《自然》杂志上表明P7采用了一种相当复杂的六聚体组装方式之前，P7的名字被分配给p7，这意味着一个简单的孔。这种新的结构暗示了一种由丙型肝炎病毒开发的新的通道机制，但目前，钙离子选择性和离子传导的结构基础尚不清楚。已有研究表明，阻断p7通道活性可减少感染性病毒后代的产生，并且已鉴定出几种化合物。这些药物相互作用应该为合理的药物开发提供有用的信息，但没有关于这些化合物在p7通道中如何和在哪里发挥作用的信息。此外，尽管人们一致认为p7通道活性对病毒的产生很重要，但在组装和释放的哪些步骤中，通道活性所需的作用仍然不清楚。我们将利用最近的p7通道结构，并利用涉及结构生物学、通道记录、分子病毒学和药物化学的多学科方法来解决这些关键问题。在目标1中，我们将确定双细胞中p7通道的结构，确定对阳离子结合和传导至关重要的关键残基，并研究与传导机制相关的构象交换。在目标2中，我们将通过突变将p7的通道活性从其在蛋白质-蛋白质相互作用中的作用中分离出来，以了解p7通道活性在病毒感染中的作用。最后，在目标3中，我们将确定抑制剂的结合部位，描述抑制机制，并探索开发新抑制剂的策略。从拟议的研究中获得的知识可能会为开发治疗丙型肝炎病毒感染的化合物带来新的机会。