NMR Spectrometer System Optimized for Direct 13C or 15N Detection

描述(由申请人提供)：我们请求资金购买和安装核磁共振光谱仪系统，用于高灵敏地直接检测蛋白质、核酸、代谢物和天然产品中碳-13和氮-15的信号。该系统将具有多核(氢-1、氢-2、碳-13和氮-15)的能力。为了节省资金，该系统将重复使用牛津500 MHz(1H)磁铁，该磁铁可在室内使用。直接观察碳-13和氮-15已被证明是从含有顺磁性金属的蛋白质的活性部位检测核磁共振信号的唯一方法。此外，对碳-13的直接观测已显示出作为收集和指定较大蛋白质和核酸的核磁共振光谱数据的策略的巨大前景。所描述的用户项目包括蛋白质、蛋白质-蛋白质复合体、蛋白质-配体复合体、核酸、蛋白质-核酸复合体和天然产物的结构-功能研究。要研究的系统很大、很复杂，在某些情况下还含有顺磁中心。这些性质妨碍或阻碍了用标准的质子探测多核方法对其进行研究。我们提供的例子证明了直接检测方法的必要性。与目前可用的仪器相比，所要求的系统将使碳-13和氮-15直接检测实验的灵敏度提高3-4倍，从而将数据收集时间缩短10倍。该系统将使研究比目前可以考虑的更大、更难溶解和更不稳定的系统成为可能。该仪器将安装在位于麦迪逊的国家磁共振设施(NMRFAM)的太空中，已经为严格控制温度和湿度进行了配备和优化，并配备了调节电力和自备冷水系统。仪器将由在使用低温探头的核磁共振系统的操作和维护方面经验丰富的光谱学家管理，他们将提供使用仪器的培训，并与用户合作开发针对其问题的最佳数据收集方法。访问权限将根据公布的NMRFAM操作程序进行分配，并由NMRFAM的常设本地和外部咨询委员会提供监督。直接检测实验为蛋白质和核酸的结构功能研究提供了最令人兴奋的新方法之一。在所描述的系统中，有一些与人类疾病直接相关的系统：铁-硫组装(Friedreich‘s共济失调)、脂肪酸减饱和(肥胖、糖尿病)和RNA-金属相互作用(衰老、癌症)。

DESCRIPTION (provided by applicant): We request funds to purchase and install an NMR spectrometer system for highly sensitive direct detection of signals from carbon-13 and nitrogen-15 in proteins, nucleic acids, metabolites, and natural products. The system will have multinuclear (hydrogen-1, hydrogen-2, carbon-13, and nitrogen-15) capability. To save money, this system will re-use an Oxford 500 MHz (1H) magnet available in house. Direct observation of carbon-13 and nitrogen-15 has proven to be the only way to detect NMR signals from the active sites of paramagnetic metal containing proteins. In addition, direct observation of carbon-13 has shown great promise as a strategy for collecting and assigning NMR spectral data from larger proteins and nucleic acids. The user projects described cover structure-function investigations of proteins, protein-protein complexes, protein-ligand complexes, nucleic acids, protein-nucleic acid complexes, and natural products. The systems to be studied are large, complex, and in several cases contain paramagnetic centers. These properties preclude or hinder their investigation by standard proton-detected multinuclear approaches. We provide examples that demonstrate the necessity of the direct detection approach. Compared to currently available instrumentation, the requested system will increase the sensitivity of direct carbon-13 and nitrogen-15 detection experiments by a factor of 3-4 and thus decrease data collection time by a factor of 10. The system will enable the investigation of systems that are larger, less-soluble, and less-stable than can be currently considered. The instrumentation will be installed in the National Magnetic Resonance Facility at Madison (NMRFAM) in space already fitted out and optimized for tight control of temperature and humidity and with conditioned power and a captive chilled-water system. The instrumentation will be managed by spectroscopists experienced in the operation and maintenance of NMR systems with cryogenic probes, who will provide training in the use of the instrumentation and work with users to develop optimal data collection approaches specific to their problems. Access will be allocated according to published NMRFAM operating procedures, and oversight will be provided by NMRFAM's standing local and external advisory boards. Direct-detection experiments offer one the most exciting new approaches to structure-function investigations of proteins and nucleic acids. Among the systems described are ones of direct relevance to human disease: iron-sulfur assembly (Friedreich's ataxia), fatty acid desaturation (obesity, diabetes), and RNA-metal interactions (ageing, cancer).