Synthetic mutualism in engineered mutant strains as functional basis for microbial production consortia.

In nature, microorganisms often reside in symbiotic co-existence providing nutrition, stability, and protection for each partner by applying "division of labor." This principle may also be used for the overproduction of targeted compounds in bioprocesses. It requires the engineering of a synthetic co-culture with distributed tasks for each partner. Thereby, the competition on precursors, redox cofactors, and energy-which occurs in a single host-is prevented. Current applications often focus on unidirectional interactions, that is, the product of partner A is used for the completion of biosynthesis by partner B. Here, we present a synthetically engineered co-culture of two engineered mutant strains marked by the essential interaction of the partners which is achieved by implemented auxotrophies. The tryptophan auxotrophic strain ANT-3, only requiring small amounts of the aromatic amino acid, provides the auxotrophic anthranilate for the tryptophan producer TRP-3. The latter produces a surplus of tryptophan which is used to showcase the suitability of the co-culture to access related products in future applications. Co-culture characterization revealed that the microbial consortium is remarkably functionally stable for a broad range of inoculation ratios. The range of robust and functional interaction may even be extended by proper glucose feeding which was shown in a two-compartment bioreactor setting with filtrate exchange. This system even enables the use of the co-culture in a parallel two-level temperature setting which opens the door to access temperature sensitive products via heterologous production in in a continuous manner.