De novo biosynthesis of bioactive isoflavonoids by engineered yeast cell factories

TLDR

Isoflavonoids are plant natural products with significant nutraceutical, pharmaceutical, and agricultural value, yet their low natural abundance and complex structures limit access via crops or chemical synthesis, making microbial bioproduction an attractive alternative. The study aims to engineer Saccharomyces cerevisiae into a platform for efficient daidzein production, the core scaffold of isoflavonoids, and to generate bioactive glucosides from glucose. The authors rebuilt daidzein biosynthesis in yeast and, through enzyme screening, identification of rate‑limiting steps, dynamic control, substrate‑trafficking engineering, and fine‑tuning of competing pathways, increased production 94‑fold. The optimized strain yields 85.4 mg L⁻¹ daidzein, and with plant glycosyltransferases it produces 72.8 mg L⁻¹ puerarin and 73.2 mg L⁻¹ daidzin, demonstrating a viable platform for de novo isoflavonoid biosynthesis that can be extended to other complex natural products.

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.