(785d) Extending Natural Product Biosynthesis Pathways through Application of Engineered Modular E. coli Polycultures

The past few years have seen an exciting boom in research interest over the use of modular microbial co-cultures for metabolic engineering applications. The ability for these multi-strain systems to spread the metabolic burden of pathway overexpression across the members of the consortia while enabling modular-specific genetic optimization have been highlighted as a key advantages over traditional monoculture approaches. These systems have demonstrated the potential for relatively simple synthetic consortia to out perform their respective monocultures for a wide variety of target products. Until recently, efforts have primarily focused on the use of two microbial strains in co-culture. In this work we will describe our efforts to extend this technology to the use of polycultures, or three or more strains in co-culture, for the extension of natural product production pathways in E. coli. Utilizing this technique, we were able to demonstrate the de novo production of a variety of flavonoid natural products not previously obtained from monoculture efforts.

To define the production landscape for our system, we identified several key optimization parameters for the individual modules as well as the polyculture as a whole. Through the development of two new high-titer production modules for phenylpropanoic acids from tyrosine and anthocyanins from flavan-3-ols, we were able to realize the de novo production of anthocyanidin-3-O-glucosides for the first time from a recombinant cell system. In addition to describing this successful case study, we will discuss our efforts to demonstrate the feasibility of these systems for other target pathways and methods to measure population dynamics in an attempt to address the stability of these systems during extended microbial fermentations. The work presented here highlights the potential of modular microbial co-cultures to revolutionize the future of natural product production from microbial hosts while attempting to address several of the lesser understood aspects of this new and exciting field.