Bacterial Expression Core

细菌表达核心

• 批准号: 7924696
• 负责人: CHRISTOPHER P. HILL
• 金额: $ 16.81万
• 依托单位: UNIVERSITY OF UTAH
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7924696
• 关键词: Amino Acid Sequence; Back; Bacterial Proteins; Biochemical; Bioinformatics; Biology; Charge; Cloning; Core Facility; DNA Sequence; Data Base Management; Floor; Funding; Hematology; Human Resources; Mass Spectrum Analysis; Methodology; Methods; Molecular; Mutation; N-terminal; Oligonucleotides; Operative Surgical Procedures; Peptide Sequence Determination; Polymerase; Production; Proteins; Protocols documentation; Publications; Recombinants; Resources; Roentgen Rays; Services; Solubility; Standardization; Surface; TEV protease; Testing; Universities; Utah; base; cost effective; expression vector; instrumentation; member; protein expression; protein purification; protein structure; response; structural biology; trafficking

Overview The Bacterial Protein Expression Core is located on the third floor of the EEJMRB at the University of Utah In close proximity to the labs of Hill, Sundquist, and Kay. It is managed by Heidi Schubert and will be staffed by two Center-funded technicians. The facility provides efficient construction of expression vectors, testing of protein expression and solubility, protein purification, and characterization. We have established a synergistic relationship with the Utah Molecular Hematology Protein Expression Core for the cost effective production of recombinant TEV protease and Pfu polymerase, and employ services of the University of Utah core facilities in mass spectrometry, oligonucleotide synthesis, N-terminal protein sequencing, and DNA sequencing. The Bacterial Protein Expression Core facilities are integrated with operations of the Eukaryotic Protein Expression Core for bioinformatics, cloning, and protein purification, and there is also a close association with protein structure determination efforts in X-ray and Protein NMR cores. This core will therefore facilitate structural and biochemical studies on targets identified by the Center members pursuing questions of HIV/Host biology. In general, the methods used in this Core are standard, but have been adapted to maximize efficiency on a scale appropriate for the targeted, but ambitious, scope of this application. The basic instrumentation and protocols are already in place and will be expanded to match increased demand if the Center is funded. The increased throughput will be achieved by efficient database management, consolidated bioinformatics resources, use of technical staff, and standardization and refinement of protocols. The approaches are an extension of the methodology that has supported structural and biochemical studies by the Hill and Sundquist labs (see biosketches for publications). Our initial target capacity is an average of 24 new expression vectors constructed per week, although the actual rate will fluctuate in response to demand. This level of throughput will allow aggressive approaches to problems, such as optimizing N- and C-termini or making surface mutations to produce crystallizable constructs. If demand exceeds capacity, we can add personnel/instrumentation/supplies on a charge-back basis.

概述 细菌蛋白表达核心位于大学的EEJMRB的三楼 犹他州靠近希尔，桑德奎斯特和凯实验室。它由海蒂·舒伯特（Heidi Schubert）管理 由两名中央资助的技术人员组成。该设施提供了有效的表达构造 向量，蛋白质表达和溶解度的测试，蛋白质纯化和表征。我们有 与犹他州分子血液学蛋白表达核心建立了协同关系 重组TEV蛋白酶和PFU聚合酶的成本有效生产，并采用 犹他大学质谱，寡核苷酸合成，N末端蛋白质的核心设施 测序和DNA测序。细菌蛋白表达核心设施与 真核蛋白表达核心的生物信息学，克隆和蛋白质纯化的操作， X射线和蛋白质的蛋白质结构确定工作也有密切的关联 NMR核心。因此，该核心将促进对靶标的结构和生化研究 追求艾滋病毒/宿主生物学问题的中心成员。 通常，该核心中使用的方法是标准的，但已适应以最大化效率 以适合该应用程序的有针对性但雄心勃勃的范围的规模。基本 仪器和协议已经到位，如果 该中心是资助的。增加的吞吐量将通过有效的数据库管理来实现 合并的生物信息学资源，技术人员的使用以及标准化和完善 协议。这些方法是支持结构和的方法的扩展 Hill和Sundquist Labs的生化研究（有关出版物的Biosketches）。我们的最初目标 容量平均为每周构建的24个新表达矢量，尽管实际速率将 响应需求而波动。这种水平的吞吐量将使问题的积极方法， 例如优化N-和C末端或进行表面突变以产生可结晶的构建体。如果 需求超过容量，我们可以以收费的方式添加人员/仪器/用品。