Core C: Enzymology Core

核心 C：酶学核心

• 批准号: 10513682
• 负责人: CRAIG E. CAMERON
• 金额: $ 746.26万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-16 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10513682
• 关键词: Alberta province; Alphavirus; Bacteria; Binding Sites; Biochemical; Biochemistry; Biological Assay; Biophysics; COVID-19 assay; Cells; Collaborations; Communities; Complex; Coronavirus; Cryoelectron Microscopy; Development; Distant; Ebola virus; Enzymatic Biochemistry; Enzyme Inhibition; Enzyme Kinetics; Enzymes; Exhibits; Filovirus; Flavivirus; Generations; Goals; HIV-1; Insecta; International; Laboratories; Lead; Mammalian Cell; Measurement; Miniaturization; Monitor; Multienzyme Complexes; Multiprotein Complexes; Mutation; Nucleotides; Polymerase; Process; Protein Family; RNA Helicase; RNA-Directed DNA Polymerase; Records; Research Project Grants; Resistance; Resistance development; Reverse Transcriptase Inhibitors; SARS coronavirus; Structure; System; Technology; Testing; Translations; Universities; Validation; Viral; Virus; X-Ray Crystallography; antiviral drug development; biophysical techniques; density; drug development; drug discovery; high throughput screening; inhibitor; insight; interest; lead optimization; member; programs; protein expression; protein purification; protocol development; screening; single molecule; structural biology; three dimensional structure; tool; virology

ABSTRACT The overarching goal of Core C is to support READDI-AC discovery efforts in collaboration with Discovery Core B and MedChem Core D by validation, characterization, and optimization of compounds in enzyme assays and structural studies, and to provide the Research Projects with detailed mechanisms of action and mechanistic insights to mitigate potential problems with resistance conferring mutations. Enzymology Core C includes seven internationally recognized laboratories that cover complementary expertise at the interfaces of virology, biochemistry, biophysics and structural biology. Three specific aims that describe the central activities of Enzymology Core C: Aim 1: Develop assays to support hit discovery efforts by Discovery Core B. Key activities include the expression and purification of enzymes that the Research Projects and Discovery Core B have selected as targets. Enzymology Core C will support the development of protocols for the expression and purification of new enzyme targets and their corresponding assays to monitor activity and inhibition. The translation of enzyme assays into high density formats to facilitate high throughput screening by Discovery Core B is a second responsibility. Aim 2. Structural Biology to guide hit and lead optimization. Enzymology Core C will focus on solving 3D structures of polymerases and RNA helicases that are currently not available to the scientific community. Priority will also be given to solving the structures of flavivirus and alphavirus polymerases in complex with the nucleotide and non-nucleotide analog inhibitors to provide detailed information on the inhibitor binding sites. Aim 3. Support the Projects in detailed mechanism of action studies for lead compounds. Functional assays will be employed to assess the inhibitory activity against the biologically relevant enzyme target, such as measurements of the half maximal inhibitory concentration to determine compound potency. Promising hits and leads will also be tested against homologous enzymes within the protein family as well as distant enzymes to assess breadth and the potential for off-target activity. Enzymology Core C will provide an array of state-of-the-art biochemical/biophysical approaches to provide MedChem Core D and the Projects with information on the mechanisms associated with inhibition and resistance.

摘要 Core C的首要目标是与Discovery Core协作支持READDI-AC发现工作 B和MedChem Core D通过在酶测定中验证、表征和优化化合物， 结构研究，并为研究项目提供详细的作用机制和机制 洞察力，以减轻潜在的问题与耐药性赋予突变。 酶学核心C包括七个国际公认的实验室，涵盖互补的专业知识 在病毒学、生物化学、生物物理学和结构生物学的交界处。三个具体目标描述了 酶学核心C的核心活动： 目标1：开发检测试剂盒以支持Discovery Core B的命中发现工作。 主要活动包括表达和纯化酶，研究项目和发现 核心B被选为目标。Enzymology Core C将支持开发用于 新酶靶的表达和纯化及其相应的检测以监测活性， 抑制作用将酶测定转化为高密度形式以促进高通量筛选， 发现核心B是第二责任。 目标2.结构生物学指导命中和领先优化。 Enzymology Core C将专注于解决聚合酶和RNA解旋酶的3D结构， 这是科学界所不具备的。还将优先解决黄病毒的结构， 与核苷酸和非核苷酸类似物抑制剂复合的甲病毒聚合酶，以提供详细的 关于抑制剂结合位点的信息。 目标3.支持项目对先导化合物进行详细的作用机制研究。 将采用功能测定来评估对生物学相关酶的抑制活性 目标，例如测量半最大抑制浓度以确定化合物效力。 有希望的命中和线索也将针对蛋白质家族内的同源酶以及 远距离酶来评估宽度和脱靶活性的潜力。酶学核心C将提供一个 一系列最先进的生化/生物物理方法，为MedChem Core D和项目提供 与抑制和抗性相关的机制的信息。