Understanding and Controlling Viral Membrane Fusion Proteins

了解和控制病毒膜融合蛋白

• 批准号: 1264506
• 负责人: Eric Boder
• 金额: $ 32万
• 依托单位: University of Tennessee Knoxville
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-05-15 至 2017-04-30
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1264506&HistoricalAwards=false
• 关键词: Understanding; Controlling; Viral; Membrane; Fusion

1264506 Boder, Eric Enveloped viruses (those surrounded by a lipid bilayer membrane,) are covered with numerous copies of a unique molecular machine, the envelope fusion protein, that is responsible for targeting the virus to a specific cell, sensing the appropriate environment for infection, and efficiently transporting the viral genetic payload across the barrier of the target cell membrane -- the key functions in all forms of delivery. The investigators recently discovered a novel intermolecular communication phenomenon associated with the activity of influenza hemagglutinin, leading to "prion-like" autocatalysis of a major conformational change involved in driving membrane fusion. It is proposed to study the mechanism of autocatalytic cross-talk and to apply directed evolution to gain control over the biosensing capabilities of this prototypical viral fusion protein, potentiating use of these engineered proteins in lipid vesicle-based platforms for a variety of applications. Using conditions that perturb cell membranes and in vitro reconstitution of hemagglutinin into synthetic lipid bilayers, the hypothesis will be tested that modulation of local membrane properties plays a role in autocatalytic communication. Both structure-directed and random mutagenesis will be applied, the latter combined with library screening and high-throughput sequencing, to map the mutational plasticity of hemagglutinin. Finally, the knowledge of mutationally tolerant structural sites in hemagglutinin will be used to design libraries in directed evolution experiments, aiming to isolate novel mutants capable of activation and fusogenic activity in response to novel stimuli. This project will enhance the understanding of molecular mechanisms critical to the life cycle of many viruses, with possible implications in designing future therapeutic interventions. In addition, new molecules to be generated in this project and the tools and methods used to identify them may be immediately useful in creating new components to be used in cell-specific drug or gene delivery. Other applications for such molecular components, such as development of membrane-based "smart" materials that respond to a variety of stimuli by fusing to mix their contents, may be possible, as well. Finally, the student personnel educated within this researchproject will be uniquely equipped with interdisciplinary skills and will be poised to lead STEM education and practice of the future.This award by the Biotechnology, Biochemical, and Biomass Engineering Program of the CBET Division is co-funded by the Biomaterials Program of the Division of Materials Research.

小行星1264506 包膜病毒（被脂质双层膜包围的病毒）覆盖着许多独特分子机器的拷贝，包膜融合蛋白，负责将病毒靶向特定细胞，感知感染的适当环境，并有效地将病毒遗传有效载荷穿过靶细胞膜的屏障-所有形式的递送中的关键功能。研究人员最近发现了一种与流感血凝素活性相关的新的分子间通讯现象，导致参与驱动膜融合的主要构象变化的“朊病毒样”自催化。建议研究自催化串扰的机制，并应用定向进化来获得对这种原型病毒融合蛋白的生物传感能力的控制，从而增强这些工程蛋白在基于脂质囊泡的平台中用于各种应用的用途。使用的条件，扰乱细胞膜和在体外重建的血凝素合成脂质双层，假设将被测试，局部膜特性的调制中发挥作用的自催化通信。结构定向和随机诱变都将被应用，后者与文库筛选和高通量测序相结合，以绘制血凝素的突变可塑性。最后，在血凝素的突变耐受性结构位点的知识将被用来设计定向进化实验中的文库，旨在分离能够激活和融合活性，以响应新的刺激的新的突变体。该项目将提高对许多病毒生命周期至关重要的分子机制的理解，并可能对设计未来的治疗干预措施产生影响。此外，在该项目中产生的新分子以及用于识别它们的工具和方法可能会立即用于创建用于细胞特异性药物或基因递送的新组件。这种分子成分的其他应用，如开发基于膜的“智能”材料，通过融合混合其内容物来响应各种刺激，也是可能的。最后，在这个研究项目中接受教育的学生人员将具有独特的跨学科技能，并将随时准备引领未来的STEM教育和实践。CBET部门的生物技术，生物化学和生物质工程项目的这个奖项由材料研究部门的生物材料项目共同资助。