Integrated state-of-the-art LC-MS/NMR system for the University of Wisconsin

说明（由申请人提供）：申请资金用于购买集成液相色谱、质谱仪和核磁共振（LC-MS/NMR）系统。该系统将存放在位于威斯康星大学麦迪逊分校生物化学系的麦迪逊国家磁共振设施（NMRFAM）中。该系统将安装在一个温度和湿度严格控制的房间里，所有需要的公用设施都已经到位。该系统将用于支持正在进行的NIH支持的四个广泛领域的研究项目：（1）代谢物混合物的分析（代谢组学）。(2)药物-受体（配体-蛋白质）相互作用的筛选。(3)通过NMR筛选蛋白质和核酸样品，以在各种条件下正确折叠，并通过MS进行质量控制（正确的身份，标记和纯度）。(4)仅少量可用的天然产物的结构解析。LC-MS/NMR将具有在这个校园或目前美国任何其他学术设施中无与伦比的独特能力。该系统将具有自动化、多样品操作和样品温度控制。它将能够获得样品等分试样（未分馏或LC馏分）的ESI/APCI Q-TOF MS或MS/MS数据，这些数据也可以通过NMR光谱进行研究。核磁共振波谱仪将包括一个600兆赫屏蔽磁铁，一个多核控制台和一个低温平台。用户可以选择两个三重共振（1H或13 C观察，13 C或15 N去耦，2 H锁定）低温探头：对于足够的样品，5 mm探头将提供最高的灵敏度（300？L样品管）;对于样品有限的情况下，1.7 mm探头将提供上级性能（30？试管中的L样品）。这种核磁共振系统将能够经济地筛选蛋白质和核酸，并研究只能少量获得的天然产物。该系统将有一个固相萃取附件，能够将样品浓缩10倍。液体处理系统将样品转移到MS系统或NMR管中。恒温进样器将处理试管中NMR样品的自动筛选。该系统将有一个自动识别和定量代谢物和相关生物合成途径的软件系统。NMRFAM的工作人员将维护仪器，协助进行研究，并培训希望亲身体验如何安全和正确使用系统的用户。将征收使用费以收回运营开支。时间安排将由现有的网络系统进行，监督将由现有的地方和外部咨询委员会提供。公共卫生相关性：该仪器支持的研究预计将有益于公共卫生，因为它旨在调查衰老和各种疾病状态的代谢标志物，并确定候选新药和抗生素的天然产物的结构。该仪器将能够更全面地研究与人类健康有关的蛋白质和核酸的结构，以及它们如何与药物和调节其活性的天然分子相互作用。

DESCRIPTION (provided by applicant): Funds are requested for the purchase of an integrated liquid-chromatography, mass spectrometer, and nuclear magnetic resonance (LC-MS/NMR) system. This system will be housed in the National Magnetic Resonance Facility at Madison (NMRFAM) located in the Biochemistry Department at the University of Wisconsin-Madison. The system will be installed in a room with tightly controlled temperature and humidity and with all needed utilities already in place. The system will be used to support ongoing, NIH-supported, research projects in four broad areas: (1) Analysis of metabolite mixtures (metabolomics). (2) Screening of drug-receptor (ligand- protein) interactions. (3) Screening of protein and nucleic acid samples by NMR for proper folding under various conditions and by MS for quality control (correct identity, labeling, and purity). (4) Elucidation of structures of natural products available only in small quantities. The LC-MS/NMR will have unique capabilities unmatched on this campus or at currently any other academic facility in the US. The system will have automated, multiple sample operation with temperature control of samples. It will have the ability to acquire ESI/APCI Q-TOF MS or MS/MS data on aliquots of samples (unfractionated or LC fraction) that can be investigated also by NMR spectroscopy. The NMR spectrometer will consist of a 600 MHz shielded magnet, a multinuclear console, and a cryogenic platform. Users will have the choice of two triple-resonance (1H or 13C observe, 13C or 15N decoupling, 2H lock) cryogenic probes: for adequate samples, a 5 mm probe will provide the highest sensitivity (300 ?L samples in tubes); for sample-limited situations, a 1.7 mm probe will provide superior performance (30 ?L samples in tubes). This NMR system will enable the economical screening of proteins and nucleic acids and investigations of natural products obtainable only in small quantities. The system will have a solid-phase extraction accessory capable of concentrating samples by up to 10-fold. A liquids handling system will transfer samples to the MS system or NMR tubes. A thermostatted sample changer will handle the automated screening of NMR samples in tubes. The system will have a software system for automated identification and quantification of metabolites and related biosynthetic pathways. NMRFAM staff members will maintain the instrumentation, assist in performing studies, and train users who desire hands-on experience how to use the system safely and properly. User fees will be levied to recover operating expenses. Scheduling will be by an existing web-based system, and oversight will be provided by existing local and external advisory committees. PUBLIC HEALTH RELEVANCE: The research supported by this instrumentation is expected to benefit public health because it aims to investigate metabolic markers of aging and various disease states and to determine the structures of natural products that are candidate new drugs and antibiotics. The instrumentation will enable more complete investigations of the structures of proteins and nucleic acids related to human health and how they interact with drugs and naturally occurring molecules that modulate their activity.