Distributing a metabolic pathway among a microbial consortium enhances production of natural products

Abstract

Metabolic engineering of microorganisms such as Escherichia coli and Saccharomyces cerevisiae to produce high-value natural metabolites is often done through functional reconstitution of long metabolic pathways. Problems arise when parts of pathways require specialized environments or compartments for optimal function. Here we solve this problem through co-culture of engineered organisms, each of which contains the part of the pathway that it is best suited to hosting. In one example, we divided the synthetic pathway for the acetylated diol paclitaxel precursor into two modules, expressed in either S. cerevisiae or E. coli, neither of which can produce the paclitaxel precursor on their own. Stable co-culture in the same bioreactor was achieved by designing a mutualistic relationship between the two species in which a metabolic intermediate produced by E. coli was used and functionalized by yeast. This synthetic consortium produced 33 mg/L oxygenated taxanes, including a monoacetylated dioxygenated taxane. The same method was also used to produce tanshinone precursors and functionalized sesquiterpenes.

Engineering Escherichia coli coculture systems for the production of biochemical products

“…However, previous research on microbial consortia was primarily concerned with the study of mixed population stability and dynamic interactions (7)(8)(9)(10)(11), although a few recent studies have reported the engineering of microbial consortia for utilization of simple sugars to make small molecules of central carbon metabolism, such as ethanol and lactate (12,13). Progress was made recently, when a full n-butanol pathway was expressed in two separate E. coli cells to achieve higher production (14) and when a bacterium-yeast coculture was used to address the difficulties of functional reconstitution of a pathway involving prokaryotic and eukaryotic enzymes in a consortium, and thus improved production of complex pharmaceutical molecules (15). Here, we expand the generality of the coculture engineering by demonstrating that microbial cocultures can also be engineered to overcome more universal challenges in metabolic engineering, including high-level intermediate secretion and low-efficiency sugar mixture utilization.…”

De novo Biosynthesis of “Non‐Natural” Thaxtomin Phytotoxins

“…The rise of synthetic biology has provided access to larger synthetic DNA constructs, rapid DNA capture,7, 8, 9 editing,10, 11 assembly,12, 13 and other advances 14, 15. The prospect of using these new tools to assemble de novo biosynthetic pathways in well‐characterized heterologous host strains16, 17 for diversification and optimization of natural products, derived from less tractable microorganisms, is an attractive goal.…”

Metabolic Engineering of Microorganisms for the Production of Natural Compounds

“…After optimization of culture conditions, this microbial consortium produced 33 mg L −1 of oxygenated taxanes. [99] …”

“…[99] Taxadiene is synthesized from GGPP by taxadiene synthase, and the following reaction for oxygenated taxanes biosynthesis requires cytochrome P450, which is hardly expressed in E. coli. Therefore, E. coli was used for overproducing taxadiene, since it was more feasible to direct the metabolic flux toward the product, and yeast was engineered for converting taxadiene to oxygenated taxanes for its higher capability of expressing eukaryotic P450 enzymes.…”

“…[99,166] Several coculture methods have been developed including the coculture of multiple strains utilizing different substrates or producing different products. Meanwhile, one recently emerged technique called modular cocultivation indicates that the whole pathway can be divided and introduced into each strain working as a module.…”