CAREER: A Multidisciplinary Approach for the Discovery and Characterization of Hormone Inducers of Natural Product Biosynthetic Gene Clusters

Purdue University, West Lafayette IN

Investigators

Abstract

With the support of the Chemistry of Life Processes program in the Division of Chemistry, Elizabeth Parkinson of Purdue University is studying the regulation of Streptomyces natural product biosynthesis by hormones and their repressors. Natural products from the soil-dwelling bacteria Streptomyces are a bountiful source of medicines, agricultural products, and chemical tools for manipulating and studying biological processes. While genomics data suggests that hundreds of thousands of novel natural products likely to have interesting bioactivities remain to be discovered from Streptomyces, many are not produced when grown in the laboratory. This is likely because the bacteria are not exposed to the same chemical stimuli that they encounter in the soil. Unfortunately, the chemical signals that regulate the production of the majority of these natural products remain a mystery. Understanding these signals is critical to accessing these natural products and ultimately maximizing the natural product potential from Streptomyces. The proposed experimentation is directed at developing improved methods for accessing novel hormones and determining the natural products they regulate. The project will also implement an education plan to improve rural student access to science, science identity and interest. Specifically, a project-based learning laboratory for rural high schools focused on isolation of antibiotic-producing bacteria from their backyard will be developed. Additionally, a modified version of the lab appropriate for younger students to perform at rural children’s museums will be generated. γ-Butyrolactones and butenolides are two of the known classes of Streptomyces hormones that regulate natural product biosynthesis. Over half of Streptomyces strains are predicted to have genes regulated by these hormones, but few examples are known because 1) hormones are produced at very low quantities and 2) no rapid, efficient assays exist to identify them. The central hypothesis of this research project is that repressors in sequence similarity networks cluster based on the structure of their hormone ligands and their DNA-binding domains. To test this, divergent synthetic routes that allow easy access to an expanded library of hormones will be developed. This library includes hormones predicted based on the hormone biosynthetic gene clusters that exist within the genomic neighborhood of the receptors. High throughput methods (i.e. GFP-reporter assays and DNA affinity purification sequencing) will then be utilized to explore ligand and binding-site specificity for a library of receptors. These assays are expected to provide valuable insight into the mechanisms by which hormones affect natural product biosynthesis. They will also allow for testing of the hypothesis that sequence similarity networks of receptors are predictive of ligand and DNA-binding specificity. If correct, this will enable development of a predictive model for hormone activation that will likely permit the discovery of novel natural products. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.