Implementing Innovative Approaches to Access the Hidden Metabolomes of Bacteria

实施创新方法来获取细菌隐藏的代谢组

基本信息

批准号：
8955195
负责人：
Mohammad R Seyedsayamdost
金额：
$ 234.49万
依托单位：
PRINCETON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2015
资助国家：
美国
起止时间：
2015-09-30 至 2020-06-30
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): Microbial natural products, also referred to as secondary metabolites, have traditionally served as a significant source of drug leads, especially in the development of antibiotics. Recent years, however, have witnessed a decline in the rate of discovery of natural products-based anti-infectives. The growing resistance of bacterial pathogens to our existing arsenal of drugs coupled with our inability to generate novel antibiotics between the 1960s and the early 2000s have catapulted infectious diseases to one of the key health challenges of the 21st century. In this application, the roadblocks for finding new bioactive molecules from microbial sources are discussed and innovative, multi-disciplinary approaches for their rapid discovery are proposed. The major challenge in finding new natural products is that most microbial biosynthetic gene clusters - the sets of genes that synthesize complex secondary metabolites from simple building blocks - are not expressed under normal growth conditions. These are referred to as `silent' or `cryptic' gene clusters and methods that activate them would have a major impact on drug discovery. A second challenge is that activation of silent gene clusters and structural elucidation of the resulting products are low-throughput processes by existing approaches. To address these challenges, we propose several new and high-throughput strategies for awakening silent gene clusters. In one approach, we propose the use of multiple genetic reporters to provide a simultaneous read-out for the expression of 4-5 silent gene clusters. High-throughput screening of small molecule libraries is then used to identify elicitors or activators of these clusters. In a second strategy,we propose methods based on mass spectrometry (MS) as a read-out for activation of cryptic clusters, while elicitor screening would provide the requisite small molecules. Both approaches, multiple genetic reporters and MS-based methods, enable high-throughput discovery of cryptic metabolites. To accelerate identification and structural elucidation of these molecules, we propose application of HPLC-MS- based differential metabolomics in concert with MS/MS networking schemes. Finally, the underlying regulatory pathways that turn on silent clusters in response to exogenous small molecules will also be examined. Together, the approaches above aim to rapidly discover both elicitors and products of cryptic gene clusters, while allowing an understanding of the endogenous regulatory pathways that mediate these processes. Subsequent bioassays carried out in house and by submission to the NIH and the NCI will assess the potential therapeutic utility of bacterial cryptic metabolites.

描述（由适用提供）：传统上，微生物天然产品（也称为二级代谢产物）传统上是药物铅的重要来源，尤其是在抗生素的开发中。然而，近年来，基于天然产品的反感染剂的发现率下降了。细菌病原体对我们现有药物库的日益增长的耐药性，加上我们无法产生新型抗生素从1960年代到2000年代初期，将传染病推动了21世纪的关键健康挑战之一。在此应用中，讨论了从微生物来源找到新的生物活性分子的障碍，并提出了创新的，创新的多学科方法来快速发现。寻找新天然产物的主要挑战是，大多数微生物生物合成基因簇 - 在正常生长条件下未表达从简单构建基块中合成复杂二级代谢物的基因集。这些被称为“沉默”或“神秘”基因簇和激活它们的方法将对药物发现产生重大影响。第二个挑战是，无声基因簇的激活和所得产品的结构阐明是通过现有方法的低通量过程。为了应对这些挑战，我们提出了几种新的和高通量的策略，以唤醒无声的基因簇。在一种方法中，我们建议使用多个遗传记者为4-5个无声基因簇的表达提供简单的读数。然后，使用小分子库的高通量筛选来识别这些簇的引起剂或激活因子。在第二种策略中，我们提出了基于质谱法（MS）的方法，作为激活加密群集的读取方法，而INICOTIT筛选将提供必要的小分子。两种方法，多个遗传记者和基于MS的方法都可以对加密代谢产物进行高通量发现。为了加速这些分子的识别和结构阐明，我们建议与MS/MS网络方案一起使用基于HPLC-MS的差异代谢组学。最后，还将检查基本的调节途径，这些途径也将被检查以响应外源小分子而打开无声簇。同时，上面的方法旨在快速发现引起群和隐源性簇的产物，同时允许了解介导这些过程的内源性调节途径。随后在内部进行的生物测定法，并通过服从NIH，NCI将评估细菌加密代谢物的潜在治疗用途。

项目成果

期刊论文数量（18）

专著数量（0）

科研奖励数量（0）

会议论文数量（0）

专利数量（4）

Thailandenes, Cryptic Polyene Natural Products Isolated from Burkholderia thailandensis Using Phenotype-Guided Transposon Mutagenesis.

DOI：
10.1021/acschembio.9b00883
发表时间：
2020-05-15
期刊：
ACS CHEMICAL BIOLOGY
影响因子：
4
作者：
Park, Jong-Duk;Moon, Kyuho;Miller, Cheryl;Rose, Jessica;Xu, Fei;Ebmeier, Christopher C.;Jacobsen, Eremy R.;Mao, Dainan;Old, William M.;DeShazer, David;Seyedsayamdost, Mohammad R.
通讯作者：
Seyedsayamdost, Mohammad R.

The Polyene Natural Product Thailandamide A Inhibits Fatty Acid Biosynthesis in Gram-Positive and Gram-Negative Bacteria.