Method for Awakening Silent Gene Clusters in Bacteria and Discovery of Cryptic Metabolites

摘要

The majority of clinically used antibiotics and anticancer agents are derived from bacterial small molecules. These molecules are produced by dedicated biosynthetic gene clusters, sets of genes that are responsible for the step-wise generation of the target small molecule. Recent investigations have indicated, to the surprise of many experts, that the majority of these biosynthetic genes are inactive or ‘silent’ for unknown reasons. Thus under typical bacterial culturing conditions, these genes are not expressed and consequently the bioactive small molecule products are not synthesized. Disclosed is a method for high throughput screening of elicitors of cryptic metabolites, a method for producing cryptic metabolites, and a new family of cryptic metabolites, the acybolins, as well as their complete structural elucidation.

主权项

1. A high-throughput screening method for discovering an agent which is able to activate silent bacterial gene clusters, comprising the steps of:
providing at least a first and second bacterial cell, wherein the bacterial cells contains at least one gene cluster that is silent or lowly-expressed and wherein the gene cluster is genetically modified to include at least one reporter gene within the gene cluster; exposing the second bacterial cell to a test compound; measuring the expression of at least one reporter gene in the first bacterial cell; measuring the expression of at least one reporter gene in the second bacterial cell; and identifying activation of gene clusters by determining whether at least one reporter gene has been expressed by the second bacterial cell by a statistically significant amount greater than is expressed by the first bacterial cell.