Biosynthesis of antifungal nucleoside antibiotics

项目摘要/摘要 本设备补充项目旨在购买一台高效液相色谱联用质谱计 (LCMS)发现、生物合成和开发抗真菌核苷所必需的仪器 抗生素。亲本R01旨在了解肽核苷(PN)抗真菌抗生素的生物合成 抗“谷热”病原等多种病原真菌并发现新的病原 利用这一知识的抗真菌药物。对于这项研究来说，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传统的 反相柱最常用于LC MS。为了克服这一挑战，我们建议购买 生物相容(无金属)超高效液相色谱(UHPLC)装置与 四极飞行时间(QTOF)质谱仪。生物相容的超高效液相将允许使用各种亲水性 相互作用色谱柱(HILIC)用于分析极性代谢物。QTOF MS探测器将 允许对复杂的代谢物混合物进行以发现为导向(无针对性)的分析。拟议的系统将使 在监测MS色谱图的同时对分析物进行纯化，这是其他LC MS所不能做到的 杜克大学提供的仪器。此外，拥有专门用于准备和分析的仪器 亲水性化合物将显著缩短我们分析的周转时间，从2-3周缩短到一周 天。因此，购买拟议的LCMS将提供我们目前无法提供的独特能力 但对于母项目中提出的核苷类抗生素的成功表征是必不可少的。这个 所提出的研究具有重要的意义，因为它将为未来的化学酶制剂或 基因组挖掘发现新型核苷类抗真菌药物。

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.