Construction of oleanolic acid-producing Saccharomyces cerevisiae cells.
10.19540/j.cnki.cjcmm.20250216.105
- Author:
Yue ZHANG
1
;
Xue-Mi HAO
2
;
Cai-Xia WANG
2
;
Long-Shan ZHAO
3
Author Information
1. School of Pharmacy, Baotou Medical College, Inner Mongolia University of Science and Technology Baotou 014040, China Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences Beijing 100700, China.
2. Institute of Chinese Materia Medica, China Academy of Chinese Medical Sciences Beijing 100700, China.
3. School of Pharmacy, Baotou Medical College, Inner Mongolia University of Science and Technology Baotou 014040, China School of Pharmacy, Shenyang Pharmaceutical University Shenyang 110016, China.
- Publication Type:Journal Article
- Keywords:
PPP approach;
Saccharomyces cerevisiae;
oleanane triterpenoid saponin;
oleanolic acid;
synthetic biology
- MeSH:
Oleanolic Acid/biosynthesis*;
Saccharomyces cerevisiae/metabolism*;
Industrial Microbiology;
Microorganisms, Genetically-Modified/metabolism*;
Plants/enzymology*;
Fermentation;
Metabolic Engineering
- From:
China Journal of Chinese Materia Medica
2025;50(9):2365-2372
- CountryChina
- Language:Chinese
-
Abstract:
In this study, Saccharomyces cerevisiae R0 was used as the chassis cell to synthesize oleanolic acid from scratch through the heterologous expression of β-amyrin synthase(β-AS) from Glycyrrhiza uralensis, cytochrome P450 enzyme CYP716A154 from Catharanthus roseus, and cytochrome P450 reductase AtCPR from Arabidopsis thaliana. The engineered strain R1 achieved shake flask titres of 5.19 mg·L~(-1). By overexpressing enzymes in the pentose phosphate pathway(PPP)(ZWF1, GND1, TKL1, and TAL), the NADH kinase gene in the mitochondrial matrix(POS5), truncated 3-hydroxy-3-methylglutaryl-CoA reductase(tPgHMGR1) from Panax ginseng, and farnesyl diphosphate synthase gene(SmFPS) from Salvia miltiorrhiza, the precursor supply and intracellular reduced nicotinamide adenine dinucleotide phosphate(NADPH) supply were enhanced, resulting in an 11.4-fold increase in squalene yield and a 3.6-fold increase in oleanolic acid yield. Subsequently, increasing the copy number of the heterologous genes tPgHMGR1, β-AS, CYP716A154, and AtCPR promoted the metabolic flow towards the final product, oleanolic acid, and increased the yield by three times. Shake flask fermentation data showed that, by increasing the copy number, precursor supply, and intracellular NADPH supply, the final engineered strain R3 could achieve an oleanolic acid yield of 53.96 mg·L~(-1), which was 10 times higher than that of the control strain R1. This study not only laid the foundation for the green biosynthesis of oleanolic acid but also provided a reference for metabolic engineering research on other pentacyclic triterpenoids in S. cerevisiae.
