Biosynthesis of antifungal nucleoside antibiotics

项目总结/摘要 本设备补充项目拟采购一台液相色谱-质谱联用仪 （LCMS）仪器是发现、生物合成和开发抗真菌核苷的关键 抗生素亲本R 01旨在了解肽基核苷（PN）抗真菌抗生素的生物合成 对多种病原性真菌如“山谷热”的病原体具有活性，并发现新的 抗真菌药物使用这些知识。对于这项研究，最重要的技术挑战之一是 亲水性核苷化合物的表征，因为这些化合物不与常规的核苷化合物结合， 最常用于LCMS的反相柱。为了克服这一挑战，我们建议购买 生物相容性（无金属）超高效液相色谱（UHPLC）单元与 四极杆飞行时间（QTOF）MS装置。生物相容性UHPLC将允许使用各种亲水性的 用于分析极性代谢物的相互作用色谱（HILIC）柱。QTOF MS检测器将 允许对复杂的代谢物混合物进行发现导向的（非目标的）分析。拟议的系统将使 分析物的纯化，同时监测MS色谱图，这是不可能的其他LCMS 杜克大学提供的仪器。此外，具有专用于制备和分析 亲水性化合物将显著改善我们分析的周转时间，从2-3周缩短到 天因此，购买拟议的LCMS将提供我们目前无法提供的独特功能 但是对于母项目中提出的核苷抗生素的成功表征是必不可少的。的 拟议的研究是重要的，因为它将为未来的化学酶制剂提供基础， 新核苷抗真菌药的基因组挖掘发现。

Project Summary/Abstract This equipment supplement project aims to purchase a liquid chromatography coupled mass spectrometry (LCMS) instrument essential for the discovery, biosynthesis, and development of antifungal nucleoside antibiotics. The parent R01 aims to understand the biosynthesis of peptidyl nucleoside (PN) antifungal antibiotics active against multiple pathogenic fungi such as the causal agent of "Valley Fever" and to discover novel antifungal agents using this knowledge. For this study, one of the most significant technical challenges is the characterization of hydrophilic nucleoside compounds because these compounds do not bind to the conventional reverse-phase columns most frequently used for LCMS. To overcome this challenge, we propose the purchase of a biocompatible (metal-free) ultra-high-performance liquid chromatography (UHPLC) unit coupled with a quadrupole time of flight (QTOF) MS device. Biocompatible UHPLC will allow the use of various hydrophilic interaction chromatography (HILIC) columns for the analysis of polar metabolites. The QTOF MS detector will allow discovery-oriented (untargeted) analyses of complex metabolite mixtures. The proposed system will enable the purification of the analytes while monitoring MS chromatograms, which is impossible with other LCMS instruments available at Duke. Also, having an instrument dedicated to the preparation and analysis of hydrophilic compounds will significantly improve the turnaround time of our analyses from 2-3 weeks to within a day. Therefore, the purchase of the proposed LCMS will provide unique capabilities currently unavailable to us but essential for the successful characterization of nucleoside antibiotics proposed in the parent project. The proposed research is significant because it will provide a basis for the future chemoenzymatic preparation or genome mining discovery of novel nucleoside antifungals.