A tripartite microbial co-culture system for de novo biosynthesis of diverse plant phenylpropanoids

Abstract

Plant-derived phenylpropanoids, in particular phenylpropenes, have diverse industrial applications ranging from flavors and fragrances to polymers and pharmaceuticals. Heterologous biosynthesis of these products has the potential to address low, seasonally dependent yields hindering ease of widespread manufacturing. However, previous efforts have been hindered by the inherent pathway promiscuity and the microbial toxicity of key pathway intermediates. Here, in this study, we establish the propensity of a tripartite microbial co-culture to overcome these limitations and demonstrate to our knowledge the first reported de novo phenylpropene production from simple sugar starting materials. After initially designing the system to accumulate eugenol, the platform modularity and downstream enzyme promiscuity was leveraged to quickly create avenues for hydroxychavicol and chavicol production. The consortia was found to be compatible with Engineered Living Material production platforms that allow for reusable, cold-chain-independent distributed manufacturing. This work lays the foundation for further deployment of modular microbial approaches to produce plant secondary metabolites.

Simple phenylpropanoids: recent advances in biological activities, biosynthetic pathways, and microbial production

“…In addition, mutualistic consortia also endow microorganisms with higher synthetic titres through unique abilities, such as balancing metabolic flow, relieving metabolic burden, optimising resource utilisation, enriching the cellular environment and cofactors, and adapting to fluctuating environments. Considering these advantages, microbial consortium strategies have been extended to construct bacterial, yeast, and yeast systems for the synthesis of terpenoids, 214 phenylpropanoids, 215,216 and polyketones, 217 and alkaloids. 218 However, most are simply mixed cultures formed by changing the initial inoculation ratio, 219 which makes it difficult to form a stable community, and this instability is further amplified in large-scale fermentation.…”

“…Efforts to construct a mutualistic relationship for the members of microbial consortia to form stable mutualistic consortia is a viable approach to improve these strategies. 216,220…”

Effect of Silica Xerogel Functionalization on Intensification of Rindera graeca Transgenic Roots Proliferation and Boosting Naphthoquinone Production

“…One of the practical bioengineering techniques widely recognized as a high-potential solution for the intensification of secondary plant metabolite production is in situ extraction [16][17][18][19]. Moreover, such a technique also provides a favorable microenvironment for the de novo production of secondary metabolites, which do not occur in the natural environment [20][21][22][23].…”

Improving Production of Coniferyl Alcohol in Escherichia coli by Regulating the Multiple Cofactors and Efflux of Intermediates

“…7−10 Since the content of free coniferyl alcohol in plants is relatively low, 11 microbial cell factories offer a promising approach to produce coniferyl alcohol and its high value-added derivatives. 4,12,13 Biosynthesis of coniferyl alcohol involves many promiscuous enzymes (Figure 1), including 4hydroxyphenylacetate 3-monooxygenase (HpaBC), 4-hydroxycinnamate:CoA ligase (4CL), cinnamoyl-CoA reductase (CCR), and cinnamyl alcohol dehydrogenase (CAD). 1 Through the screening of key enzymes, comparisons of fermentation medium, and the addition of ascorbic acid, engineered Escherichia coli could produce 187.7 mg/L coniferyl alcohol in a 5-L bioreactor.…”

“…5,11,19 However, eugenol could only be commercially obtained from plant extraction and is not cheap. 13 Hence, there is an urgent need to construct an economical coniferyl alcohol biosynthetic pathway. Some phenolic acids and monolignols can inhibit microbial growth, which is a limiting factor for the microbial synthesis of coniferyl alcohol, 20 and screening and the expression of efflux proteins are key to aromatic synthesis.…”