Innovative Technologies for Metabolite Profiling and Natural Products Discovery

描述(申请人提供)：自然界的小分子宇宙提供了非常丰富的结构多样性、物理化学特征和生物活性。对自然产生的小分子的研究可分为两类：(1)代谢组学，即研究细胞和组织的小分子含量，以确定(病理)重要的生理生化途径；(2)天然产物的发现，或鉴定具有生物活性的化合物。最终，对代谢物和天然产物的研究取决于能够从复杂的生物样品中有效分离和浓缩它们的方法。尽管作出了相当大的努力，但这些化合物的浓缩和表征的全球战略仍然难以捉摸。在这项应用中，我们建议开发一套创新的技术，以促进任何物理化学类别的小分子的浓缩。 指导K99阶段：该计划的博士后阶段(目标1)将在斯克里普斯研究所本杰明·克拉瓦特教授的指导下进行。这一阶段的目标是开发一种适用于代谢组谱的化学选择性标记和浓缩策略。这项培训将提供质谱学、结构测定和细胞培养技术方面的专业知识。所获得的知识将为目标2和目标3以及向独立职业生涯过渡奠定基础。 独立的R00阶段：AIM 2将专注于应用AIM 1中开发的策略，结合基于活动的蛋白质组和基因组技术，组装有助于细菌持久性的生化网络。医院病原体的一种持久菌株铜绿假单胞菌将通过与Floyd Romesberg教授的合作进行检查。AIM 3将扩展小分子标记策略，以创建一种浓缩生物活性天然产品的技术。这种方法将有助于检测使用当前纯化方法往往无法获得的天然产物，并能够以前所未有的广度和深度探索大自然的小分子谱系。 相关性：该计划有可能通过开发有助于了解疾病发病机制的技术以及发现治疗剂来改善公共健康。我们将应用这些方法来研究铜绿假单胞菌的持久性和致病性，以及新的抗菌药的鉴定。

DESCRIPTION (provided by applicant): Nature's small molecule universe offers a remarkably rich assortment of structural diversity, physicochemical characteristics, and bioactivity. The study of naturally occurring small molecules can be divided into two categories: (1) metabolomics, or study of the small molecule content of cells and tissues to identify (patho)physiologically important biochemical pathways, and (2) natural products discovery, or the identification of compounds possessing biological activity. Ultimately, the investigation of metabolites and natural products depends upon methods that enable their effective separation and enrichment from complex biological samples. Despite considerable effort, global strategies for the enrichment and characterization of these compounds have remained elusive. In this application, we propose to develop an innovative set of technologies to facilitate the enrichment of small molecules of any physico-chemical class. Mentored K99 Phase: The postdoctoral phase (aim 1) of the proposal will be carried out under the mentorship of Prof. Benjamin Cravatt at the Scripps Research Institute. The goal of this phase is to develop a chemo-selective tagging and enrichment strategy applicable to metabolomic profiling. This training will provide expertise in mass spectrometry, structure determination, and cell culture techniques. The acquired knowledge will provide the foundation for aims 2 and 3 and transition to an independent career. Independent R00 Phase: Aim 2 will focus on application of the strategy developed in aim 1 in combination with activity-based proteomic and genomic technologies to assemble biochemical networks that contribute to bacterial persistence. A persistent strain of the hospital pathogen, Pseudomonas aeruginosa, will be examined through collaboration with Prof. Floyd Romesberg. Aim 3 will expand on the small-molecule tagging strategy to create a technology for the enrichment of bioactive natural products. This method will facilitate detection of natural products that are often inaccessible using current purification methods and enable exploration of nature's small molecule repertoire with unprecedented scope and depth. Relevance: This program has the potential to improve public health through development of technologies that will assist in understanding disease pathogenesis, as well as discovery of therapeutic agents. We will apply these methods to the study of persistence and pathogenicity in Pseudomonas aeruginosa and the identification of novel antimicrobials.