Mechanisms of Enterovirus Entry

肠道病毒进入机制

• 批准号: 8960335
• 负责人: Susan Hafenstein
• 金额: $ 39.67万
• 依托单位: PENNSYLVANIA STATE UNIV HERSHEY MED CTR
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-11-01 至 2019-10-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8960335
• 关键词: 3-Dimensional; Acute; Acute Disease; Affinity; Aseptic Meningitis; Binding; Biological Assay; CAR receptor; CD55 Antigens; Capsid; Cell membrane; Cells; Cleaved cell; Collaborations; Communicable Diseases; Complex; Coxsackie B Viruses; Coxsackie Viruses; Cryoelectron Microscopy; Crystallography; Data; Encephalitis; Enterovirus; Epithelial; Event; Genome; Goals; Health; Heating; Hour; Human; In Situ; Infection; Insulin-Dependent Diabetes Mellitus; Lead; Length; Lipid Bilayers; Location; Mass Spectrum Analysis; Membrane; Methods; Modeling; Molecular; Motion; Myocarditis; Pancreatitis; Paralysed; Pediatric Hospitals; Peptides; Pharmaceutical Preparations; Philadelphia; Physiological; Play; Proteins; Proteolysis; RNA; Resolution; Role; Signal Transduction; Staging; Stimulus; Structure; Surface Plasmon Resonance; Testing; Tight Junctions; Transmembrane Domain; Viral Genome; Virus; Virus Receptors; adenovirus receptor; base; density; design; endosome membrane; factor A; improved; innovation; insulin dependent diabetes mellitus onset; interest; nanodisk; novel; particle; pathogen; receptor; receptor binding; recombinant virus; reconstruction; research study; respiratory; trafficking

DESCRIPTION (provided by applicant): Using structural approaches (cryo-electron microscopy and x-ray crystallography), we will investigate the mechanisms used by coxsackievirus B3 (CVB3) to enter host cells by engaging two receptors (decay accelerating factor and coxsackievirus and adenovirus receptor). CVB3 is a human pathogen that causes myocarditis, pancreatitis, and has been implicated in the onset of juvenile diabetes mellitus. Our proposed studies of CVB3 entry are a continuation of a productive collaboration with Dr. Jeffery Bergelson, Division of Infectious Diseases at Children's Hospital at Philadelphia. A combination of traditional and innovative methods will be used to visualize both receptor binding and the resulting conformational changes that lead to entry and uncoating. Of particular interest is a novel entry intermediate that has never been observed (Aim 1). This particle has been isolated with fingerlike density projections extruding from the capsid, and these densities are hypothesized to be peptides used to anchor the particle to the host membrane. We will investigate the capability of this new particle (and other entry intermediates) to bind receptors and infect the host. In addition to conventional structural studies made with purified virus and recombinant receptor proteins in solution (Aim 2), the full-length receptor will be anchored onto a lipid bilayer disc to form virus-receptor complexes for an in situ asymmetric study (Aim 3). This approach simulates the physiological binding event, in which the virus interacts with receptors at a single, focused region on its capsid. Preliminary results with this asymmetric approach suggest the virus behaves differently when interacting with a few membrane-bound receptors compared to many soluble receptors. Specifically our in situ A-particle retains an intact genome (as the A-particle does in an infection) and undergoes different protein rearrangements compared to particles that are treated with soluble receptors or heat. The proposed study is directed at understanding the structural alterations that an enterovirus capsid must undergo to uncoat after successfully finding and attaching to the receptor in situ.

描述（由申请人提供）：使用结构方法（冷冻电子显微镜和X射线晶体学），我们将研究Coxsackievivirus B3（CVB3）使用的机制，通过参与两个受体（衰减加速因子和Coxsackievievirus和Coxsackivivirus和Adenovirus受体）进入宿主细胞。 CVB3是一种引起心肌炎，胰腺炎的人类病原体，并与青少年糖尿病的发作有关。我们对CVB3进入的拟议研究是与费城儿童医院的传染病科Jeffery Bergelson博士进行了富有成效的合作。传统和创新方法的结合将用于可视化受体结合以及导致进入和脱落的结果构象变化。特别有趣的是一个从未观察到的新型进入中间体（AIM 1）。该粒子已被带有帽子挤出的指形的密度突显分离，并且这些密度被认为是将粒子固定在宿主膜上的肽。我们将研究这种新粒子（和其他进入中间体）结合受体并感染宿主的能力。除了在溶液中用纯化的病毒和重组受体蛋白进行的常规结构研究（AIM 2），全长受体还将锚定在脂质双层盘上，以形成病毒受体复合物，以进行原位不对称研究（AIM 3）。这 方法模拟了生理结合事件，其中该病毒与单个聚焦区域的受体在其衣壳上相互作用。这种不对称方法的初步结果表明，与许多可溶性受体相比，与少数膜结合的受体相互作用时，病毒的行为不同。具体来说，我们的原位A粒子保留了完整的基因组（如A颗粒在感染中所做的那样），并且与用可溶受体或热处理的颗粒相比，经历了不同的蛋白质重排。拟议的研究旨在理解肠内capsID在成功找到并附着在原位受体之后必须进行的结构改变。