Metabolic Engineering of Yarrowia lipolytica for De Novo Biosynthesis of Umbelliferone
- 1Shanghai Key Laboratory of Bio-Energy Crops, School of Life Sciences, Shanghai University, Shanghai 200444, China
Abstract
Umbelliferone, a key precursor for the biosynthesis of high-value furanocoumarin derivatives, is produced via the shikimate pathway. The oleaginous yeast Yarrowia lipolytica possesses an inherently robust shikimate pathway; however, de novo biosynthesis of umbelliferone in this microbial chassis has not been realized to date. Low intracellular pools of p-coumaric acid and suboptimal catalytic activity of downstream hydroxylation and cyclization enzymes represent the major bottlenecks limiting efficient umbelliferone production. In this study, we systematically screened five PAL/TAL variants, three 4CL isoforms, and two C2′H enzymes derived from diverse plant and microbial sources. The bifunctional enzyme RtPAL-TAL exhibited the highest productivity of p-coumaric acid, while the combination of At4CL3 and RgC2′H achieved the maximum umbelliferone accumulation. Expression of feedback-insensitive shikimate pathway mutant enzymes effectively relieved intrinsic pathway feedback inhibition, resulting in a 10.2-fold improvement in p-coumaric acid titer. All synthetic biosynthetic gene cassettes were stably integrated into the Y. lipolytica genome to generate the engineered strain YL-PU, which yielded 3.29 mg/L umbelliferone in shake-flask cultures. Further fed-batch fermentation in a 5 L bioreactor increased the umbelliferone titer to 125.52 mg/L at 84 h of cultivation. This work establishes the first de novo umbelliferone biosynthesis pathway in Y. lipolytica and identifies that C2′H represents a major candidate bottleneck under the tested configuration, in agreement with earlier findings. These findings provide a robust foundation for further metabolic engineering of the Y. lipolytica chassis to enable high-titer production of coumarins and other phenylpropanoid natural products.
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BibTeX
@article{lu2026metabolic,
title = {{Metabolic Engineering of Yarrowia lipolytica for De Novo Biosynthesis of Umbelliferone}},
author = {Fengliu Lu and Ying Luo and Zhen Lei and Changfu Li and Yansheng Zhang},
journal = {Biomolecules},
year = {2026},
month = oct,
doi = {10.3390/biom16101450},
url = {https://www.mdpi.com/2218-273X/16/10/1450},
note = {CC BY 4.0},
}
RIS
TY - JOUR
TI - Metabolic Engineering of Yarrowia lipolytica for De Novo Biosynthesis of Umbelliferone
AU - Fengliu Lu
AU - Ying Luo
AU - Zhen Lei
AU - Changfu Li
AU - Yansheng Zhang
PY - 2026
DA - 2026/10/04
JO - Biomolecules
DO - 10.3390/biom16101450
UR - https://www.mdpi.com/2218-273X/16/10/1450
AB - Umbelliferone, a key precursor for the biosynthesis of high-value furanocoumarin derivatives, is produced via the shikimate pathway. The oleaginous yeast Yarrowia lipolytica possesses an inherently robust shikimate pathway; however, de novo biosynthesis of umbelliferone in this microbial chassis has not been realized to date. Low intracellular pools of p-coumaric acid and suboptimal catalytic activity of downstream hydroxylation and cyclization enzymes represent the major bottlenecks limiting efficient umbelliferone production. In this study, we systematically screened five PAL/TAL variants, three 4CL isoforms, and two C2′H enzymes derived from diverse plant and microbial sources. The bifunctional enzyme RtPAL-TAL exhibited the highest productivity of p-coumaric acid, while the combination of At4CL3 and RgC2′H achieved the maximum umbelliferone accumulation. Expression of feedback-insensitive shikimate pathway mutant enzymes effectively relieved intrinsic pathway feedback inhibition, resulting in a 10.2-fold improvement in p-coumaric acid titer. All synthetic biosynthetic gene cassettes were stably integrated into the Y. lipolytica genome to generate the engineered strain YL-PU, which yielded 3.29 mg/L umbelliferone in shake-flask cultures. Further fed-batch fermentation in a 5 L bioreactor increased the umbelliferone titer to 125.52 mg/L at 84 h of cultivation. This work establishes the first de novo umbelliferone biosynthesis pathway in Y. lipolytica and identifies that C2′H represents a major candidate bottleneck under the tested configuration, in agreement with earlier findings. These findings provide a robust foundation for further metabolic engineering of the Y. lipolytica chassis to enable high-titer production of coumarins and other phenylpropanoid natural products.
SN - 2218-273X
ER -