DEVELOPMENT OF INDUCTION AND STRUCTURAL ELUCIDATION METHODS FOR THE DISCOVERY OF CRYPTIC BACTERIAL NATURAL PRODUCTS

Abstract

Natural products encompass a diverse array of small molecules and harbor exquisite therapeutic properties and intriguing ecological functions. Recent advancements in genome sequencing have unveiled a large collection of silent biosynthetic gene clusters within microorganisms, the activation of which promises to unearth novel natural products. This thesis is dedicated to developing innovative methods to trigger the production of these ‘cryptic’ natural products in bacteria and elucidating the structure, biosynthesis, and bioactivity of the newly discovered molecules. Chapter 2 reports the induction of three cryptic peptidic natural products from Streptomyces rimosus, the renowned oxytetracycline producer, using the MALDI-MS guided high-throughput elicitor screening approach. Elucidating the chemical structures and the biosynthetic origins of these molecules reveals momomycin, a novel cyclic peptide harboring back-bone modifications and non-canonical amino acids, which exhibits potent antiproliferative activities against cancer cells. Furthermore, investigations into the activation mechanism of the momomycin gene cluster shed light on the regulatory intricacies involving induction of natural product biosynthesis. Chapter 3 explores the discovery of eight novel and structurally distinctive alkaloid natural products from the bacterial symbiont, Phaeobacter inhibens, facilitated by supplementing algal metabolites to cell cultures. These newly identified molecules displayed potent algaecidal activities, underscoring the bacterium’s ability to convert algal host precursors into complex cytotoxins during the parasitic phase of the symbiotic interaction. Structure elucidation of three compounds, produced in minute quantities, employs the emerging microcrystal diffraction technique of cryogenic electron microscopy. In summary, this thesis showcases the application of chemical genetics and culture modulation methods to activate silent biosynthetic gene clusters. The uncovered natural products demonstrate significant structural complexity and novelty, which are essential for equipping them with promising pharmaceutical values and relevant ecological functions. This work contributes to our understanding of natural product biosynthesis, silent gene cluster regulation, and chemical-mediated microbial interactions.