Catalytic activity of the two-component flavin-dependent monooxygenase from Pseudomonas aeruginosa toward cinnamic acid derivatives

Furuya, Toshiki; Kino, Kuniki

doi:10.1007/s00253-013-4958-y

Cited by 61 publications

(61 citation statements)

References 52 publications

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“…This stepwise approach was tested due to the possible toxicity of p-coumaric and ferulic acids to the cells [12,[18][19][20][21][22], and because we found that this strategy improved the production of caffeic acid in our previous work (manuscript submitted). However, in the current study the two-step feeding was shown to be counterproductive, i.e.…”

Section: Discussionmentioning

confidence: 99%

Production of curcuminoids from tyrosine by a metabolically engineered Escherichia coli using caffeic acid as an intermediate

Rodrigues

Araújo

Prather

et al. 2015

Biotechnology Journal

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Curcuminoids are phenylpropanoids with high pharmaceutical potential. Herein, we report an engineered artificial pathway in Escherichia coli to produce natural curcuminoids through caffeic acid. Arabidopsis thaliana 4-coumaroyl-CoA ligase (4CL1) and Curcuma longa diketide-CoA synthase (DCS) and curcumin synthase (CURS1) were used to produce curcuminoids and 70 mg/L of curcumin was obtained from ferulic acid. Bisdemethoxycurcumin and demethoxycurcumin were also produced, but in lower concentrations, by feeding p-coumaric acid or a mixture of p-coumaric acid and ferulic acid, respectively. Additionally, curcuminoids were produced from tyrosine through the caffeic acid pathway. To produce caffeic acid, tyrosine ammonia lyase (TAL) from Rhodotorula glutinis and 4-coumarate 3-hydroxylase (C3H) from Saccharothrix espanaensis were used. Caffeoyl-CoA 3-O-methyltransferase (CCoAOMT) from Medicago sativa was used to convert caffeoyl-CoA to feruloyl-CoA.Using caffeic acid, p-coumaric acid or tyrosine as a substrate, 3.9 mg/L, 0.3 mg/L and 0.2 mg/L of curcumin were produced, respectively. This is the first time DCS and CURS1 were used in vivo to produce curcuminoids and that curcumin was produced by feeding tyrosine. We have shown that curcumin can be produced using a pathway through caffeic acid. This alternative pathway represents a step forward in the heterologous production of curcumin using E. coli.

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Section: Discussionmentioning

confidence: 99%

Production of curcuminoids from tyrosine by a metabolically engineered Escherichia coli using caffeic acid as an intermediate

Rodrigues

Araújo

Prather

et al. 2015

Biotechnology Journal

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“…The pETDhpaBC plasmid carrying the hpaB and hpaC genes, which was previously constructed, 17) was introduced into Escherichia coli BL21 Star (DE3) cells (Invitrogen, Carlsbad, CA, USA). The transformed E. coli cells were cultivated at 25°C in Luria-Bertani medium, which contained (per liter) Bacto tryptone (10 g), Bacto yeast extract (5 g), and NaCl (10 g, pH 7.0), supplemented with ampicillin (100 μg mL −1 ).…”

Section: Methodsmentioning

confidence: 99%

“…17,18) The reaction mixture (100 μL) was acidified by the addition of HCl (pH 2-3), and methanol (500 μL) and water (400 μL) were added to the reaction mixture. After vigorous shaking and centrifugation, the resulting supernatant (10 μL) was injected into the HPLC system.…”

Section: Product Analysismentioning

confidence: 99%

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Biocatalytic synthesis of 3,4,5,3′,5′-pentahydroxy-trans-stilbene from piceatannol by two-component flavin-dependent monooxygenase HpaBC

Furuya

Sai

Kino

2016

Bioscience, Biotechnology, and Biochemistry

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HpaBC monooxygenase was previously reported to hydroxylate resveratrol to piceatannol. In this article, we report a novel catalytic activity of HpaBC for the synthesis of a pentahydroxylated stilbene. When Escherichia coli cells expressing HpaBC were incubated with resveratrol, the resulting piceatannol was further converted to a new product. This product was identified by mass spectrometry and NMR spectroscopy as a 5-hydroxylated piceatannol, 3,4,5,3',5'-pentahydroxy-trans-stilbene (PHS), which is a reportedly valuable biologically active stilbene derivative. We attempted to produce PHS from piceatannol on a flask scale. After examining the effects of detergents and buffers on PHS production, E. coli cells expressing HpaBC efficiently hydroxylated piceatannol to PHS in a reaction mixture containing 1.5% (v/v) Tween 80 and 100 mM 3-morpholinopropanesulfonic acid-NaOH buffer at pH 7.5. Under the optimized conditions, the whole cells regioselectively hydroxylated piceatannol, and the production of PHS reached 6.9 mM (1.8 g L(-1)) in 48 h.

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“…Tyrosine ammonia lyase and 4-coumarate 3-hydroxylase, encoded by sam8 and sam5, from Saccharothrix espanaensis were used to convert L-tyrosine caffeic acid 4b. 17 pHydroxyphenylacetate (HPA) 3-hydroxylase (HPAH) from E. coli 18 and Pseudomonas aeruginosa 19 can catalyze the hydroxylation of 2-(4-hydroxyphenyl)acetic acid (4-HPA) 2a and pcoumaric acid 4a to 2-(3,4-dihydroxyphenyl)acetic acid (3, 2b and caffeic acid 4b, respectively. However, these enzymatic systems cannot efficiently prepare trihydroxyphenolic acids.…”

Section: ■ Introductionmentioning

confidence: 99%

p-Hydroxyphenylacetate 3-Hydroxylase as a Biocatalyst for the Synthesis of Trihydroxyphenolic Acids

et al. 2015

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Trihydroxyphenolic acids such as 3,4,5-trihydroxycinnamic acid (3,4,5-THCA) 4c and 2-(3,4,5-trihydroxyphenyl)acetic acid (3,4,5-THPA) 2c are strong antioxidants that are potentially useful as medicinal agents. Our results show that p-hydroxyphenylacetate (HPA) 3-hydroxylase (HPAH) from Acinetobacter baumannii can catalyze the syntheses of 3,4,5-THPA 2c and 3,4,5-THCA 4c from 4-HPA 2a and p-coumaric acid 4a, respectively. The wildtype HPAH can convert 4-HPA 2a completely into 3,4,5-THPA 2c within 100 min (total turnover number (TTN) of 100). However, the wild-type enzyme cannot efficiently synthesize 3,4,5-THCA 4c. To improve the efficiency, the oxygenase component of HPAH (C 2 ) was rationally engineered in order to maximize the conversion of p-coumaric acid 4a to 3,4,5-THCA 4c. Results from site-directed mutagenesis studies showed that Y398S is significantly more effective than the wildtype enzyme for the synthesis of 3,4,5-THCA 4c; it can catalyze the complete bioconversion of p-coumaric acid 4a to 3,4,5-THCA 4c within 180 min (TTN ∼ 23 at 180 min). The yield and stability of 3,4,5-THPA 2c and 3,4,5-THCA 4c were significantly improved in the presence of ascorbic acid. Thermostability studies showed that the wild-type C 2 was very stable and remained active after incubation at 30, 35, and 40°C for 24 h. Y398S was moderately stable because its activity was retained for 24 h at 30°C and for 15 h at 35°C. Transient kinetic studies using stopped-flow spectrophotometry indicated that the key improvement in the reaction of Y398S with p-coumaric acid 4a lies within the protein−ligand interaction. Y398S binds to pcoumaric acid 4a with higher affinity than the wild-type enzyme, resulting in a shift in equilibrium toward favoring the productive coupling path instead of the path leading to wasteful flavin oxidation.

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Catalytic activity of the two-component flavin-dependent monooxygenase from Pseudomonas aeruginosa toward cinnamic acid derivatives

Cited by 61 publications

References 52 publications

Production of curcuminoids from tyrosine by a metabolically engineered Escherichia coli using caffeic acid as an intermediate

Production of curcuminoids from tyrosine by a metabolically engineered Escherichia coli using caffeic acid as an intermediate

Biocatalytic synthesis of 3,4,5,3′,5′-pentahydroxy-trans-stilbene from piceatannol by two-component flavin-dependent monooxygenase HpaBC

p-Hydroxyphenylacetate 3-Hydroxylase as a Biocatalyst for the Synthesis of Trihydroxyphenolic Acids

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