Abstract
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 for the biosynthesis of caffeic acid via episomal plasmid-mediated expression of pathway genes. However, the production was far from satisfactory and even relied on the addition of precursor. In this study, we first established a controllable and stable caffeic acid pathway by employing a modified GAL regulatory system to control the genome-integrated pathway genes in S. cerevisiae and realized biosynthesis of 222.7 mg/L caffeic acid. Combinatorial engineering strategies including eliminating the tyrosine-induced feedback inhibition, deleting genes involved in competing pathways, and overexpressing rate-limiting enzymes led to about 2.6-fold improvement in the caffeic acid production, reaching up to 569.0 mg/L in shake-flask cultures. To our knowledge, this is the highest ever reported titer of caffeic acid synthesized by engineered yeast. This work showed the prospect for microbial biosynthesis of caffeic acid and laid the foundation for constructing biosynthetic pathways of its derived metabolites.
Key points
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Genomic integration of ORgTAL, OHpaB, and HpaC for caffeic acid production in yeast.
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Feedback inhibition elimination and Aro10 deletion improved caffeic acid production.
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The highest ever reported titer (569.0 mg/L) of caffeic acid synthesized by yeast.
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Data availability
All experimental data and strains constructed in this study will be made available from the corresponding author upon reasonable request from readers.
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Funding
This work was financially supported by the National Natural Science Foundation of China (Grant No. 32001032), Natural Science Foundation of Jiangsu Province, China (Grant No. BK20200946), China Postdoctoral Science Foundation (Grant No. 2020M671614), Natural Science Research of Jiangsu Higher Education Institutions of China (20KJD416003), the Innovative and Entrepreneurial Talent of Jiangsu Province, and “Lvyang Jinfeng” Talent Attracting Plan of Yangzhou.
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PPZ conceived and designed the experiments. PPZ and CLY performed the experiments. BS, YD, and NNX assisted in experiments. PPZ wrote and edited the manuscript. LDY contributed materials and revised the manuscript. All authors read and approved the final manuscript.
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Zhou, P., Yue, C., Shen, B. et al. Metabolic engineering of Saccharomyces cerevisiae for enhanced production of caffeic acid. Appl Microbiol Biotechnol 105, 5809–5819 (2021). https://doi.org/10.1007/s00253-021-11445-1
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DOI: https://doi.org/10.1007/s00253-021-11445-1