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Biological synthesis of coumarins in Escherichia coli.

Yang SM, Shim GY, Kim BG, Ahn JH - Microb. Cell Fact. (2015)

Bottom Line: The solubility of F6'H was critical for the final yield.Umbelliferone and esculetin were also synthesized from glucose using engineered E. coli strains.The final yields of umbelliferone and esculetin were 66.1 and 61.4 mg/L, respectively.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioscience and Biotechnology, Bio/Molecular Informatics Center, Konkuk University, 120 Neungdong-ro, Gwangjin-gu, Seoul, 143-701, Korea. lcanldyl@nate.com.

ABSTRACT

Background: Coumarins are a major group of plant secondary metabolites that serves as defense compounds against pathogens. Although coumarins can be obtained from diverse plant sources, the use of microorganisms to synthesize them could be an alternative way to supply building blocks for the synthesis of diverse coumarin derivatives.

Results: Constructs harboring two genes, F6'H (encoding feruloyl CoA 6' hydroxylase) and 4CL (encoding 4-coumarate CoA:ligase), were manipulated to increase the productivity of coumarins. Escherichia coli expressing the two genes was cultured in medium supplemented with hydroxycinnamic acids (HCs) including p-coumaric acid, caffeic acid, and ferulic acid, resulting in the synthesis of the corresponding coumarins, umbelliferone, esculetin, and scopoletin. Cell concentration and initial substrate feeding concentration were optimized. In addition, umbelliferone, and esculetin were synthesized from glucose by using a ybgC deletion mutant and co-expressing tyrosine ammonia lyase and other genes involved in the tyrosine biosynthesis pathway.

Conclusions: To produce coumarin derivatives (umbelliferone, scopoletin, and esculetin) in E. coli, several constructs containing F6'H and 4CL were made, and their ability to synthesize coumarin derivatives was tested. The solubility of F6'H was critical for the final yield. After optimization, 82.9 mg/L of umbelliferone, 79.5 mg/L of scopoletin, and 52.3 mg/L of esculetin were biosynthesized from the corresponding HCs, respectively in E. coli. Umbelliferone and esculetin were also synthesized from glucose using engineered E. coli strains. The final yields of umbelliferone and esculetin were 66.1 and 61.4 mg/L, respectively.

No MeSH data available.


Related in: MedlinePlus

Production of scopoletin by feeding ferulic acid to E. coli strain B-CM1, A: ferulic acid standard (F), B: scopoletin standard (S), C: reaction products, P1 and P2, D: Mass spectrum of reaction product P1.
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Fig2: Production of scopoletin by feeding ferulic acid to E. coli strain B-CM1, A: ferulic acid standard (F), B: scopoletin standard (S), C: reaction products, P1 and P2, D: Mass spectrum of reaction product P1.

Mentions: Coumarin is synthesized from hydroxycinnamic acid by the products of two genes, 4CL and F6′H. Genes from Oryza sativa (Os4CL) and I. batatas (IbF6′H2) were cloned and transformed into E. coli. The E. coli transformant (B-CM1 in Table 1) was fed with ferulic acid. The reaction product was analyzed using high performance liquid chromatography (HPLC). As shown in Figure 2C, the culture filtrate from B-CM1 showed a new peak that had the same retention time as a standard of scopoletin. The molecular mass of the reaction product was 192-Da (g/mol) (Figure 2D), which corresponds to the predicted molecular mass of scopoletin. B-CM1 produced 3.5 mg/L scopoletin.Table 1


Biological synthesis of coumarins in Escherichia coli.

Yang SM, Shim GY, Kim BG, Ahn JH - Microb. Cell Fact. (2015)

Production of scopoletin by feeding ferulic acid to E. coli strain B-CM1, A: ferulic acid standard (F), B: scopoletin standard (S), C: reaction products, P1 and P2, D: Mass spectrum of reaction product P1.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4419511&req=5

Fig2: Production of scopoletin by feeding ferulic acid to E. coli strain B-CM1, A: ferulic acid standard (F), B: scopoletin standard (S), C: reaction products, P1 and P2, D: Mass spectrum of reaction product P1.
Mentions: Coumarin is synthesized from hydroxycinnamic acid by the products of two genes, 4CL and F6′H. Genes from Oryza sativa (Os4CL) and I. batatas (IbF6′H2) were cloned and transformed into E. coli. The E. coli transformant (B-CM1 in Table 1) was fed with ferulic acid. The reaction product was analyzed using high performance liquid chromatography (HPLC). As shown in Figure 2C, the culture filtrate from B-CM1 showed a new peak that had the same retention time as a standard of scopoletin. The molecular mass of the reaction product was 192-Da (g/mol) (Figure 2D), which corresponds to the predicted molecular mass of scopoletin. B-CM1 produced 3.5 mg/L scopoletin.Table 1

Bottom Line: The solubility of F6'H was critical for the final yield.Umbelliferone and esculetin were also synthesized from glucose using engineered E. coli strains.The final yields of umbelliferone and esculetin were 66.1 and 61.4 mg/L, respectively.

View Article: PubMed Central - PubMed

Affiliation: Department of Bioscience and Biotechnology, Bio/Molecular Informatics Center, Konkuk University, 120 Neungdong-ro, Gwangjin-gu, Seoul, 143-701, Korea. lcanldyl@nate.com.

ABSTRACT

Background: Coumarins are a major group of plant secondary metabolites that serves as defense compounds against pathogens. Although coumarins can be obtained from diverse plant sources, the use of microorganisms to synthesize them could be an alternative way to supply building blocks for the synthesis of diverse coumarin derivatives.

Results: Constructs harboring two genes, F6'H (encoding feruloyl CoA 6' hydroxylase) and 4CL (encoding 4-coumarate CoA:ligase), were manipulated to increase the productivity of coumarins. Escherichia coli expressing the two genes was cultured in medium supplemented with hydroxycinnamic acids (HCs) including p-coumaric acid, caffeic acid, and ferulic acid, resulting in the synthesis of the corresponding coumarins, umbelliferone, esculetin, and scopoletin. Cell concentration and initial substrate feeding concentration were optimized. In addition, umbelliferone, and esculetin were synthesized from glucose by using a ybgC deletion mutant and co-expressing tyrosine ammonia lyase and other genes involved in the tyrosine biosynthesis pathway.

Conclusions: To produce coumarin derivatives (umbelliferone, scopoletin, and esculetin) in E. coli, several constructs containing F6'H and 4CL were made, and their ability to synthesize coumarin derivatives was tested. The solubility of F6'H was critical for the final yield. After optimization, 82.9 mg/L of umbelliferone, 79.5 mg/L of scopoletin, and 52.3 mg/L of esculetin were biosynthesized from the corresponding HCs, respectively in E. coli. Umbelliferone and esculetin were also synthesized from glucose using engineered E. coli strains. The final yields of umbelliferone and esculetin were 66.1 and 61.4 mg/L, respectively.

No MeSH data available.


Related in: MedlinePlus