Dataset Information

Engineering Saccharomyces cerevisiae for the production of dihydroquercetin from naringenin.

ABSTRACT:

Background

Dihydroquercetin (DHQ), a powerful bioflavonoid, has a number of health-promoting qualities and shows potential as a treatment for a number of disorders. Dihydroquercetin biosynthesis is a promising solution to meet the rising demand for dihydroquercetin. However, due to the significant accumulation of eriodietyol (ERI), naringenin (NAR), dihydrokaempferol (DHK), and other metabolites, the yield of DHQ biosynthesis is low. As a result, this is the hindrance to the biosynthesis of DHQ.

Results

In this study, we proposed several strategies to enhance the product formation and reduce the metabolites in accumulation. The flavonoid 3'-hydroxylase (F3'H) and cytochrome P450 reductase from different species were co-expressed in S. cerevisiae, and the best strain expressing the P450-reductase enzyme complex (SmF3'H/ScCPR) yielded 435.7 ± 7.6 mg/L of ERI from NAR in the deepwell microplate. The product conversion rate was improved further by mutating the predicted potential ubiquitination sites to improve SmF3'H stability, resulting in a 12.8% increase in titre using the mutant SmF3'H (K290R). Besides, different F3Hs from various sources and promoters were tested for the improved DHQ production, with the best strain producing 381.2 ± 10.7 mg/L of DHQ from 1 g/L of NAR, suggesting the temporal regulation the expression of F3H is important for maximization the function of F3'H and F3H.

Conclusion

This study offers effective strategies for improving DHQ production from NAR and could be used as a reference for related research.

SUBMITTER: Yu S

PROVIDER: S-EPMC9569186 | biostudies-literature | 2022 Oct

REPOSITORIES: biostudies-literature

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Publications

Engineering Saccharomyces cerevisiae for the production of dihydroquercetin from naringenin.

Yu Shiqin S Li Mingjia M Gao Song S Zhou Jingwen J

Microbial cell factories 20221015 1

<h4>Background</h4>Dihydroquercetin (DHQ), a powerful bioflavonoid, has a number of health-promoting qualities and shows potential as a treatment for a number of disorders. Dihydroquercetin biosynthesis is a promising solution to meet the rising demand for dihydroquercetin. However, due to the significant accumulation of eriodietyol (ERI), naringenin (NAR), dihydrokaempferol (DHK), and other metabolites, the yield of DHQ biosynthesis is low. As a result, this is the hindrance to the biosynthesis ...[more]

PMID: 36243863

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Project description:BackgroundAnthocyanins are polyphenolic pigments which provide pink to blue colours in fruits and flowers. There is an increasing demand for anthocyanins, as food colorants and as health-promoting substances. Plant production of anthocyanins is often seasonal and cannot always meet demand due to low productivity and the complexity of the plant extracts. Therefore, a system of on-demand supply is useful. While a number of other (simpler) plant polyphenols have been successfully produced in the yeast Saccharomyces cerevisiae, production of anthocyanins has not yet been reported.ResultsSaccharomyces cerevisiae was engineered to produce pelargonidin 3-O-glucoside starting from glucose. Specific anthocyanin biosynthetic genes from Arabidopsis thaliana and Gerbera hybrida were introduced in a S. cerevisiae strain producing naringenin, the flavonoid precursor of anthocyanins. Upon culturing, pelargonidin and its 3-O-glucoside were detected inside the yeast cells, albeit at low concentrations. A number of related intermediates and side-products were much more abundant and were secreted into the culture medium. To optimize titers of pelargonidin 3-O-glucoside further, biosynthetic genes were stably integrated into the yeast genome, and formation of a major side-product, phloretic acid, was prevented by engineering the yeast chassis. Further engineering, by removing two glucosidases which are known to degrade pelargonidin 3-O-glucoside, did not result in higher yields of glycosylated pelargonidin. In aerated, pH controlled batch reactors, intracellular pelargonidin accumulation reached 0.01 µmol/gCDW, while kaempferol and dihydrokaempferol were effectively exported to reach extracellular concentration of 20 µM [5 mg/L] and 150 µM [44 mg/L], respectively.ConclusionThe results reported in this study demonstrate the proof-of-concept that S. cerevisiae is capable of de novo production of the anthocyanin pelargonidin 3-O-glucoside. Furthermore, while current conversion efficiencies are low, a number of clear bottlenecks have already been identified which, when overcome, have huge potential to enhance anthocyanin production efficiency. These results bode very well for the development of fermentation-based production systems for specific and individual anthocyanin molecules. Such systems have both great scientific value for identifying and characterising anthocyanin decorating enzymes as well as significant commercial potential for the production of, on-demand, pure bioactive compounds to be used in the food, health and even pharma industries.

Project description:BackgroundAnthranilate is a platform chemical used by the industry in the synthesis of a broad range of high-value products, such as dyes, perfumes and pharmaceutical compounds. Currently anthranilate is produced via chemical synthesis from non-renewable resources. Biological synthesis would allow the use of renewable carbon sources and avoid accumulation of toxic by-products. Microorganisms produce anthranilate as an intermediate in the tryptophan biosynthetic pathway. Several prokaryotic microorganisms have been engineered to overproduce anthranilate but attempts to engineer eukaryotic microorganisms for anthranilate production are scarce.ResultsWe subjected Saccharomyces cerevisiae, a widely used eukaryotic production host organism, to metabolic engineering for anthranilate production. A single gene knockout was sufficient to trigger anthranilate accumulation both in minimal and SCD media and the titer could be further improved by subsequent genomic alterations. The effects of the modifications on anthranilate production depended heavily on the growth medium used. By growing an engineered strain in SCD medium an anthranilate titer of 567.9 mg l-1 was obtained, which is the highest reported with an eukaryotic microorganism. Furthermore, the anthranilate biosynthetic pathway was extended by expression of anthranilic acid methyltransferase 1 from Medicago truncatula. When cultivated in YPD medium, this pathway extension enabled production of the grape flavor compound methyl anthranilate in S. cerevisiae at 414 mg l-1.ConclusionsIn this study we have engineered metabolism of S. cerevisiae for improved anthranilate production. The resulting strains may serve as a basis for development of efficient production host organisms for anthranilate-derived compounds. In order to demonstrate suitability of the engineered S. cerevisiae strains for production of such compounds, we successfully extended the anthranilate biosynthesis pathway to synthesis of methyl anthranilate.

Dataset Information

Engineering Saccharomyces cerevisiae for the production of dihydroquercetin from naringenin.

Background

Results

Conclusion

Publications

Engineering Saccharomyces cerevisiae for the production of dihydroquercetin from naringenin.

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