Abstract
Isoflavonoids comprise a class of plant natural products with great nutraceutical, pharmaceutical and agricultural significance. Their low abundance in nature and structural complexity however hampers access to these phytochemicals through traditional crop-based manufacturing or chemical synthesis. Microbial bioproduction therefore represents an attractive alternative. Here, we engineer the metabolism of Saccharomyces cerevisiae to become a platform for efficient production of daidzein, a core chemical scaffold for isoflavonoid biosynthesis, and demonstrate its application towards producing bioactive glucosides from glucose, following the screening-reconstruction-application engineering framework. First, we rebuild daidzein biosynthesis in yeast and its production is then improved by 94-fold through screening biosynthetic enzymes, identifying rate-limiting steps, implementing dynamic control, engineering substrate trafficking and fine-tuning competing metabolic processes. The optimized strain produces up to 85.4 mg L−1 of daidzein and introducing plant glycosyltransferases in this strain results in production of bioactive puerarin (72.8 mg L−1) and daidzin (73.2 mg L−1). Our work provides a promising step towards developing synthetic yeast cell factories for de novo biosynthesis of value-added isoflavonoids and the multi-phased framework may be extended to engineer pathways of complex natural products in other microbial hosts.
Highlights
Isoflavonoids comprise a class of plant natural products with great nutraceutical, pharmaceutical and agricultural significance
We established a yeast-based de novo production platform for the efficient production of the isoflavonoid carbon skeleton DEIN as well as the high-value glucosides PIN and DIN. This was achieved by first identifying functional biosynthetic enzymes to generate DEIN, by optimizing metabolic flux at enzyme and pathway levels to further increase DEIN titer and by introducing plant UGTs to convert DEIN to corresponding glucosides
Though S. cerevisiae is generally identified as a superior host for the functional expression of membrane-bound plant P450s over its prokaryotic counterparts, extra efforts are required to maximize their catalytic efficiency
Summary
Isoflavonoids comprise a class of plant natural products with great nutraceutical, pharmaceutical and agricultural significance. Grafting and optimizing plant biosynthetic pathways in microbial hosts is becoming a compelling route to supply plant natural products, as demonstrated by substantial biosynthesis of high-value-added alkaloids, stilbenes, and flavonoids, and terpenoids from simple sugar[11,12,13,14,15]. Based on this growing body of work, we speculated that microbial cell factories may offer the potential for the production of commercially viable isoflavonoid as well. While the reported low titers necessitate further improvement to support industrial-scale production, there have been rare efforts to engineer and optimize de novo microbial biosynthesis of isoflavones
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