UV-Based Approach for Accessing New Antibiotics

描述（由申请人提供）：该项目的目标是通过激活放线菌的沉默生物合成操纵子来发现新的抗生素。许多放线菌的全基因组测序表明，在一个给定的物种中，编码次级代谢物的操纵子比已知的化合物多10-20倍。这为获得大量未开发的新抗生素提供了一个有吸引力的机会。基因工程已被用于打开放线菌的沉默操纵子，导致次生代谢物的产生。然而，工程操纵子被激活的速度非常低，基于这种方法每年不到10种代谢产物被报道。我们推断，对体外不产生抗菌素的分离物进行诱变将使沉默操纵子免受监管约束

DESCRIPTION (provided by applicant): The goal of this project is to discover novel antibiotics by activating the silent biosynthetic operons of Actinobacteria. Whole genome sequencing of many Actinobacteria showed that there are 10-20 times more operons coding for secondary metabolites than known compounds in a given species. This opens an attractive opportunity to access a large untapped source of new antibiotics. Genetic engineering has been used to turn on silent operons in Actinobacteria, leading to production of secondary metabolites. However, the pace at which engineered operons are activated is very low, less than ten metabolites a year are being reported based on this approach. We reasoned that mutagenesis of isolates that do not produce antimicrobials in vitro will relieve silent operons from regulatory constraints, and screening will then identify the producing mutants. Our preliminary data showed that the approach works surprisingly well, turning over half of the inactive organisms into antibiotic producers. The method is scalable, and we recently identified two potentially novel antimicrobials using this approach. In Phase I, we will mutagenize/screen 2,000 inactive strains for antibiotic production, aiming to obtain new, potentially useful antimicrobials. Biological and chemical dereplication will indicate compounds with potential novelty. We will give priority to broad spectrum compounds with activity against difficult to treat gram-negative pathogens. The antimicrobials will be tested for potency, spectrum, specificity of action and cytotoxicity, and th structure of compounds that pass validation will be determined. Finding 2-3 antimicrobials with novel chemistry will serve as proof-of-principle for this approach. These findings will provide a solid basis for a large-scale drug discovery effort in Phase II. PUBLIC HEALTH RELEVANCE: The overall goal of the project is to develop a simple and efficient method to discover novel antibiotics to combat drug-resistant pathogens that pose a serious health threat in the US and around the world. Novel approaches for discovering antibiotics are desperately needed as the drug pipeline for treating these dangerous human infections continues to diminish.