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Protein Purification and Biochemistry

蛋白质纯化和生物化学

基本信息

批准号：
9233915
负责人：
JOSEPH A DUNCAN
金额：
$ 32.82万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-03-01 至 2019-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9233915
关键词：
ATP Hydrolysis Affect Affinity Antiviral Agents Baculoviridae Baculoviruses Binding Biochemical Biological Assay Cells Chromatography DNA DNA Damage Development Ensure Evaluation Excision Family Family member Goals Hepatitis A Hepatitis A Virus Herpesviridae Human Human Herpesvirus 6 Human Herpesvirus 8 Hydrolysis Immune Immune signaling In Vitro Individual Innate Immune Response Insecta Intervention Laboratories Lead Ligand Binding Liquid Chromatography Methodology Methods Molecular Natural Immunity Nucleic Acid Binding Nucleic Acids Nucleotides Pathway interactions Pattern Pattern Recognition Pattern recognition receptor Production Protein Biochemistry Proteins Protocols documentation Publishing RNA Reagent Receptor Signaling Recombinant Proteins Recombinants Research Research Personnel Resource Sharing Role Signal Pathway Signal Transduction Signaling Molecule Signaling Protein Source Standardization Structure System Systems Analysis Technology Toll-like receptors Transcript Untranslated RNA Viral Virus Virus Diseases Virus Receptors Virus Replication Work adaptive immune response base extracellular helicase novel nucleic acid detection particle pathogen protein B protein expression protein function protein purification receptor reconstitution repaired response sensor therapeutic target tool transmission process viral DNA viral RNA viral detection

项目摘要

The overall objective of this Shared Resource Core is to enhance the capacity of the project investigators of this cooperative research center to determine the novel molecular mechanisms contributing to innate immune recognition of a variety of viral pathogens by contributing to the following center goals: 1) to investigate the role novel Pattern Recognition Receptors (PRR) as authentic receptors of viral nucleic acid which affect subsequent human innate immune responses to viral pathogens and (2) to capitalize on unique biochemical capabilities to quantify the ligand-binding functions of PRRs that are technically-challenging. The core will provide center investigators with purified innate immune signaling proteins as well as specialized assays of their biochemical activities that are relevant to signal transduction. The Core will capitalize on the expertise of Dr. Duncan in recombinant protein production and characterization, biochemical reconstitution of signaling pathways, and specific work in the field of innate immune signaling in order to provide this support. The core will generate these proteins and assay the ATP binding, ATP hydrolysis, and binding to virally derived nucleic acids of each protein in order to ensure biochemically active properly functioning protein is available for the studies proposed by center investigators. The inclusion of the Protein Purification and Biochemistry Core in this U19 application will provide a common source for recombinant signaling proteins, several of which are utilized in multiple projects. Because the production and characterization of these proteins is not routine and requires specific assay technologies for nucleotide binding and hydrolysis as well as nucleic acid binding, a single Shared resource Core will allow for efficient availability these reagents to multiple laboratories and the use of robust standardized methodologies in order to assess nucleic acid sensing functions by these proteins. Overall, this core will provide tools for each project to understand the mechanisms of viral nucleic acid detection and better assess the potential for these signaling systems to be used to develop novel anti-viral strategies based on host signaling pathways.

该共享资源核心的总体目标是提高项目研究人员的能力该合作研究中心旨在确定有助于先天的新分子机制通过促进以下中心目标来免疫识别多种病毒病原体：1）研究新型模式识别受体（PRR）作为病毒核酸真实受体的作用影响随后人类对病毒病原体的先天免疫反应，以及（2）利用独特的量化 PRR 配体结合功能的生化能力在技术上具有挑战性。该核心将为中心研究人员提供纯化的先天免疫信号蛋白以及对其与信号转导相关的生化活性进行专门测定。核心将利用 Duncan 博士在重组蛋白生产和表征方面的专业知识，信号通路的生化重建以及先天免疫信号传导领域的具体工作以便提供这种支持。核心将生成这些蛋白质并测定 ATP 结合、ATP 水解，并与每种蛋白质的病毒衍生核酸结合，以确保生化活性功能正常的蛋白质可用于中心研究人员提出的研究。纳入此 U19 应用中的蛋白质纯化和生物化学核心将为重组信号蛋白，其中一些在多个项目中使用。因为生产和这些蛋白质的表征不是常规的，需要特定的核苷酸检测技术结合和水解以及核酸结合，单个共享资源核心将允许高效多个实验室可以使用这些试剂，并使用强大的标准化方法，以便评估这些蛋白质的核酸传感功能。总的来说，这个核心将为每个人提供工具项目旨在了解病毒核酸检测的机制并更好地评估这些潜力信号系统用于开发基于宿主信号通路的新型抗病毒策略。