Promoter-Library-Based Pathway Optimization for Efficient (2<i>S</i>)-Naringenin Production from <i>p</i>-Coumaric Acid in <i>Saccharomyces cerevisiae</i>

Abstract

Pathway optimization plays an important role in fine-tuning metabolic pathways. In most conditions, more than three genes are involved in the biosynthesis pathway of a specific target product. To improve the titer of products, rational regulation of a group of genes by a series of promoters with different strengths is essential. On the basis of a series of RNA-Seq data, a set of 66 native promoters was chosen to fine-tune gene expression in <i>Saccharomyces cerevisiae</i>. Promoter strength was characterized by measuring the fluorescence strength of the enhanced green fluorescent protein through fluorescence-activated cell sorting. The expressions of P_TDH1, P_PGK1, P_INO1, P_SED1, and P_CCW12 were stronger than that of P_TDH3, whereas those of another 15 promoters were stronger than that of P_TEF1. Then, 30 promoters were chosen to optimize the biosynthesis pathway of (2<i>S</i>)-naringenin from <i>p</i>-coumaric acid. With a high-throughput screening method, the highest titer of (2<i>S</i>)-naringenin in a 5 L bioreactor reached 1.21 g/L from <i>p</i>-coumaric acid, which is the highest titer according to the currently available reports.