Enhancement of β-Alanine Biosynthesis in Escherichia coli Based on Multivariate Modular Metabolic Engineering

SIMPLE SUMMARY: Pathway balancing is a common and critical challenge for the construction of microbial cell factories using metabolic engineering approaches. However, semi-rational or non-rational manipulation might lead to metabolic imbalances, which may further impact pathway efficiency, and ultimately impact the growth performance and production performance of a microbial cell factory. In this study, the multivariate modular metabolic engineering was employed to engineer the β-alanine biosynthesis pathway and keep the balance of metabolic flux among the whole metabolic network, rationally and systematically. Ultimately, 37.9 g/L β-alanine was generated in fed-batch fermentation. Novel strategies reported in this study were meaningful to the application and diffusion in β-alanine industrial production. ABSTRACT: β-alanine is widely used as an intermediate in industrial production. However, the low production of microbial cell factories limits its further application. Here, to improve the biosynthesis production of β-alanine in Escherichia coli, multivariate modular metabolic engineering was recruited to manipulate the β-alanine biosynthesis pathway through keeping the balance of metabolic flux among the whole metabolic network. The β-alanine biosynthesis pathway was separated into three modules: the β-alanine biosynthesis module, TCA module, and glycolysis module. Global regulation was performed throughout the entire β-alanine biosynthesis pathway rationally and systematically by optimizing metabolic flux, overcoming metabolic bottlenecks and weakening branch pathways. As a result, metabolic flux was channeled in the direction of β-alanine biosynthesis without huge metabolic burden, and 37.9 g/L β-alanine was generated by engineered Escherichia coli strain B0016-07 in fed-batch fermentation. This study was meaningful to the synthetic biology of β-alanine industrial production.