UCLA

Natural products, since the discovery of the first antibiotic penicillin in 1928, have been the source of and inspiration for drugs. Due the emergence of drug-resistant pathogens and discovery of drug targets in cancer biology, the need for the discovery of new bioactive natural compounds and synthesis of analogs thereof remains present. Fortuitously, the development of next-generation sequencing and tools for the heterologous expression of the biosynthetic genes for natural products has accelerated the discovery of new natural products as well as the elucidation of the respective biosynthetic pathways. This dissertation describes the use of these two emerging technologies in revealing insights in the biosynthesis of two antifungal natural compounds synthesized by filamentous fungi: griseofulvin, an antifungal drug used in treatment against dermatophytes and is in the World Health Organization List of Essential Medicine and echinocandin B, the parent compound of the frontline anti-Candidiasis drug anidulafungin.

This dissertation will first present the work in the investigation the biosynthesis of echinocandin B. Invasive candidiasis caused by opportunistic pathogenic strains of genus Candida, accounts for 17% of ICU-related infections and through understanding of the biosynthesis of echinocandin B will aid in the mutasynthesis of analogs with better efficacy than the parent compound. In this work, we characterized the key steps in the biosynthesis of the natural product. Deletion of the biosynthetic genes also allowed for the mutasynthesis of echinocandin B analogs with more potent antifungal activity and showed the potential for re-engineering of the pathway to produce “unnatural” natural compounds.

Moreover, this work also describes the full elucidation and reconstitution of the enzymatic reactions leading to the biosynthesis of griseofulvin using a combination of gene knockout and biochemical characterization of the griseofulvin biosynthetic enzymes. Among the important biosynthetic enzymes characterized in this study is the non-reducing polyketide synthase GsfA which was implicated in the biosynthesis of norlichexanthone and the cytochrome P450 GsfF which catalyzes the formation of the spirobicyclic grisan ring in griseofulvin. The culmination of this study is the demonstration of the total in vitro biosynthesis of the compound using purified griseofulvin biosynthetic enzymes.