Abstract
Synthetic consortia represent an innovative and effective platform that can significantly alleviate the metabolic burden on host organisms and enable flexible regulation of biosynthetic pathways. However, designing a stable synthetic consortium remains a significant challenge. In this study, a novel citramalate -derived pathway is first developed for 2-methylbutyric acid (2MBA) biosynthesis in an E. coli mono-culture system, achieving a titer of 678.78 ± 49.04 mg L(-1). Furthermore, it employs a CulECpy model-guided strategy to design and optimize the division of labor within E. coli synthetic consortia, predicting the optimal pathway allocation for improved 2MBA production. The best-performing consortium, using 2-keto-3-methylvalerate (KMV) as a single node, achieved 1817.03 ± 103.73 mg L(-1) of 2MBA, a 28-fold increase over the initial mono-culture strain, with the highest reported yield of 0.091 g/g glucose. This work demonstrates the effectiveness of synthetic consortia and model-guided pathway optimization for improving high-value products, a versatile strategy that can be applied to the production of other valuable metabolites.