SIG Program: AKTAxpress FPLC System for Protein Purification

描述（由申请人提供）：AKTAxpress FPLC系统用于蛋白质纯化1)项目摘要/摘要蛋白质纯化能力是结构生物学、蛋白质设计和蛋白质科学各个方面的基础。高通量自动化蛋白质纯化可以大大提高研究效率，当许多样品必须处理和多步骤纯化方案优化。加州理工学院三个领先的蛋白质研究实验室（Arnold, Rees, Tirrell）正在申请支持购买AKTAxpress FPLC（快速蛋白质液相色谱）系统。该仪器能够在无人值守的方式下进行全自动多步纯化，从而获得最高的纯度和高产量，用于晶体学和生化蛋白表征等应用。模块化设置允许不同的纯化协议同时在不同的模块上运行，使三个研究小组能够在不相互干扰的情况下纯化他们的蛋白质。该仪器将消除我们研究工作的一个非常重要的瓶颈：蛋白质纯化。目前净化系统的时间已被完全预订，并且需要大量的研究人员投入，这将与AKTAxpress一起最小化。模块化设置还允许在需要时扩展功能（增加新的净化“模块”，总共多达12个）。该仪器将支持细胞色素P450功能的结构基础研究和用于合成人类代谢物和药物铅多样化（Arnold）的工程新P450，膜蛋白晶体结构分析（Rees），以及通过掺入非天然氨基酸的工程蛋白（Tirrell）。细胞色素P450酶在人类药物代谢中起着关键作用，了解其功能对开发安全有效的治疗药物至关重要。这些酶可以用来合成真正的代谢物用于临床测试（例如毒性或增强治疗功能）。整体膜蛋白是大多数常用药物的靶点，但其结构分析仍处于起步阶段。结合非天然氨基酸的蛋白质工程为基于蛋白质的治疗和诊断以及新的研究工具提供了令人兴奋的新机会。

DESCRIPTION (provided by applicant): AKTAxpress FPLC System for Protein Purification 1) Project Summary/Abstract Protein purification capabilities underlie structural biology, protein design, and all aspects of protein science. High-throughput automated protein purification can greatly enhance research productivity when many samples must be handled and multi-step purification protocols optimized. Three leading protein research laboratories at Caltech (Arnold, Rees, Tirrell) are requesting support to purchase an AKTAxpress FPLC (Fast Protein Liquid Chromatography) system. This instrument is capable of performing fully- automated, multi-step purifications in an unattended fashion, resulting in highest possible purity and high yields for applications such as crystallography and biochemical protein characterization. The modular setup allows different purification protocols to be run simultaneously on different modules, enabling the three research groups to purify their proteins without interfering with each other. This instrument will remove a very significant bottleneck to our research efforts: protein purification. Time on current purification systems is fully booked, and requires significant researcher input that will be minimized with the AKTAxpress. The modular setup also allows expansion of capabilities when needed (with addition of new purification 'modules', for a total of up to 12). This instrument will support research into the structural basis of cytochrome P450 function and engineering new P450s for synthesis of human metabolites and drug lead diversification (Arnold), membrane protein crystallographic structure analysis (Rees), and engineering proteins through incorporation of nonnatural amino acids (Tirrell). Cytochrome P450 enzymes are key players in human drug metabolism, and understanding their function is critical to development of safe, effective therapeutics. These enzymes can be used to synthesize authentic metabolites for clinical testing (e.g. for toxicity or enhanced therapeutic function). Integral membrane proteins are targets of most popularly prescribed drugs, yet their structural analysis is still in its infancy. Protein engineering via incorporation of nonnatural amino acids offers exciting new opportunities for protein-based therapeutics and diagnostics as well as new research tools. PUBLIC HEALTH RELEVANCE: The research supported by this instrumentation grant will lead to a better understanding of drug metabolism, drug-drug interactions, and mechanisms of drug action. Protein purification is a fundamental tool in structural biology, protein design, and all aspects of protein science. Protein purification is currently a bottleneck for all three groups requesting this instrumentation, and their research productivity will be significantly enhanced by the requested protein purification system.