Date of Award

2014

Degree Type

Thesis

Degree Name

Master of Science in Pharmaceutical Sciences

Department

Biomedical and Pharmaceutical Sciences

First Advisor

David Rowley

Abstract

Natural products, and molecules derived from them, have played an extremely large role in the field of medicine and have contributed as therapies for numerous disease states. Unfortunately, the rate of discovery of medically relevant molecules from natural sources is slowing as secondary metabolite studies from microbes are increasingly yielding the re-isolation of previously known compounds. This study focused on isolating new secondary metabolites through the usage of two different techniques: (1) treatment of fungi and bacteria with epigenetic modifying compounds to induce natural product production, and (2) genome mining/bioinformatic analysis of natural product gene clusters to enhance the isolation of new compounds from previously uninvestigated species. Previous studies in the literature have shown treatment with chemical epigenetic modifiers to be successful in inducing new secondary metabolite production in fungi. At the start of my research, no natural product studies involving epigenetic modifiers had been performed in bacteria, and, to date, only one publication has appeared in the peer-reviewed literature. The approach used during my investigation was to induce new metabolite formation in the bacterium Bacillus pumilus strain RI06-95 via epigenetic modification using agents that target several epigenetic regulatory mechanisms. Reproducibility proved a difficulty as treatment of the same organism induced the production of different metabolites in repeat experiments.

The second focus of this study utilized genome mining and bioinformatic analysis of natural product gene clusters in order to isolate new secondary metabolites from organisms previously unstudied for natural product synthesis. The Secondary Metabolite Online Repository (SMOR) is a new tool that rapidly analyzes the well-curated complete microbial genome sequences located on the National Center for Biotechnology Information Genome Database and flags natural product gene clusters. Many organisms are sequenced for reasons other than determination of their biosynthetic potential, and thus have been largely overlooked for their secondary metabolite production. By using SMOR and other bioinformatic tools, this study sought to isolate natural products, particularly predicted cyclic lipopeptide antibiotics, from the potentially overlooked microorganism Bradyrhizobium sp. BTAil. Extract fractions showed bioactivity against methicillin resistant Staphylococcus aureus and structural characterization of a purified bioactive molecule indicated the potential presence of one of the predicted cyclic lipopeptides. Unfortunately due to insufficient material and long culturing times, structural elucidation of the bioactive molecules was not possible.

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