Metabolic engineering of Saccharomyces cerevisiae for enhanced production of caffeic acid

Abstract

Background As a natural phenolic acid product of plant source, caffeic acid displays diverse biological activities and acts as an important precursor for the synthesis of other valuable compounds. Limitations in chemical synthesis or plant extraction of caffeic acid trigger interest in its microbial biosynthesis. Recently, Saccharomyces cerevisiae has been reported sporadically for biosynthesis of caffeic acid via free plasmid‑mediated pathway assembly. However, the production was far from satisfactory and even relied on the addition of precursor. Results In this study, we first established a controllable caffeic acid pathway by employing a modified GAL regulatory system in S. cerevisiae and realized de novo biosynthesis of 313.8 mg/L caffeic acid from glucose. Combinatorial engineering strategies including eliminating the tyrosine-induced feedback inhibition, deleting genes involved in competing pathways and overexpressing rate-limiting enzymes led to about 2.5-fold improvement in the caffeic acid production, reaching up to 769.3 mg/L in shake-flask cultures. To our knowledge, this is the highest ever reported titer of caffeic acid de novo synthesized by engineered yeast. Conclusions Caffeic acid production in S. cerevisiae strain was successfully improved by adopting a glucose-regulated GAL system and comprehensive metabolic engineering strategies. This work showed the prospect for microbial biosynthesis of caffeic acid and laid the foundation for constructing biosynthetic pathways of its derived metabolites.

The Neurospora crassa Inducible Q System Enables Simultaneous Optogenetic Amplification and Inversion in Saccharomyces cerevisiae for Bidirectional Control of Gene Expression

“…These results imply that McLIS has lower catalytic activity than ObGES, given that the level of expression achieved with OptoINVRT7 using P GAL1-S is 2.5-fold that of P TEF1 , 21 while OptoQ-AMP4 achieves 1.4-times P TEF1 levels. Higher linalool production could likely be attained by using a mutant LIS with higher activity, 29 adding an N-terminal SKIK tag to improve expression, 30 or introducing additional copies of LIS. Nevertheless, the ability to use light to fine-tune the composition of geraniol-linalool blends in a single strain demonstrates the vast potential of combining optogenetic amplifier and inverter circuits to balance metabolic pathways, whether they carry high or low metabolic fluxes.…”