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（由申请人提供）：许多病毒基因组编码在膜中形成通道的小的整合膜蛋白，并且它们传导质子、阳离子和其他分子穿过膜屏障以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，是病毒致病性的关键，并已被追求作为抗病毒的目标。由丙型肝炎病毒（HCV）基因组编码的病毒孔蛋白p7是更重要的病毒通道之一，其已被验证为用于开发治疗HCV感染的药物的靶标。p7是一种整合的膜蛋白，其寡聚化以形成具有阳离子选择性的通道，相对于其他阳离子，对Ca 2+具有选择性;已显示其促进感染性病毒体的组装和排出。命名为p7的“viroporin”暗示了一个简单的孔，直到我们的实验室在Ouyang et al Nature 2013中显示p7采用了一种相当复杂的六聚体组装模式。这种新的结构暗示了HCV开发的一种新的通道机制，但目前，Ca 2+选择性和离子传导的结构基础尚不清楚。已经显示阻断p7通道活性减少感染性病毒后代的产生，并且已经鉴定了几种化合物。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如何以及在哪里作用于p7通道的信息。此外，尽管一致认为p7通道活性对于病毒产生是重要的，但仍不清楚在组装和释放的哪个步骤中需要通道活性的作用。我们将利用最近的p7通道结构，并利用多学科的方法，涉及结构生物学，通道记录，分子病毒学和药物化学来解决这些关键问题。在目标1中，我们将确定的p7通道的结构在bicelles，确定重要的阳离子结合和传导的关键残基，并调查相关的传导机制的构象交换。在目标2中，我们将通过诱变将p7通道活性与其在蛋白质-蛋白质相互作用中的作用分开，以了解p7通道活性在病毒感染期间的作用。最后，在目标3中，我们将鉴定抑制剂结合位点，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HCV感染的化合物带来新的机会。