De Novo Biosynthesis of <i>p</i>-Coumaric Acid in <i>E. coli</i> with a <i>trans</i>-Cinnamic Acid 4-Hydroxylase from the Amaryllidaceae Plant <i>Lycoris aurea</i>
<i>p</i>-Coumaric acid is a commercially available phenolcarboxylic acid with a great number of important applications in the nutraceutical, pharmaceutical, material and chemical industries. <i>p</i>-Coumaric acid has been biosynthesized in some engineered microbes, but the p...
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| Format: | Article |
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MDPI AG
2018-12-01
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| Series: | Molecules |
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| Online Access: | https://www.mdpi.com/1420-3049/23/12/3185 |
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| author | Yikui Li Jie Li Binbin Qian Li Cheng Sheng Xu Ren Wang |
| author_facet | Yikui Li Jie Li Binbin Qian Li Cheng Sheng Xu Ren Wang |
| author_sort | Yikui Li |
| collection | DOAJ |
| description | <i>p</i>-Coumaric acid is a commercially available phenolcarboxylic acid with a great number of important applications in the nutraceutical, pharmaceutical, material and chemical industries. <i>p</i>-Coumaric acid has been biosynthesized in some engineered microbes, but the potential of the plant CYP450-involved biosynthetic route has not investigated in <i>Escherichia coli</i>. In the present study, a novel <i>trans</i>-cinnamic acid 4-hydroxylase (C4H) encoding the <i>Lau</i>C4H gene was isolated from <i>Lycoris aurea</i> (L’ Hér.) Herb via rapid amplification of cDNA ends. Then, <i>N</i>-terminal 28 amino acids of <i>Lau</i>C4H were characterized, for the subcellular localization, at the endoplasmic reticulum membrane in protoplasts of <i>Arabidopsis thaliana</i>. In <i>E. coli</i>, <i>Lau</i>C4H without the <i>N</i>-terminal membrane anchor region was functionally expressed when fused with the redox partner of <i>A. thaliana</i> cytochrome P450 enzyme (CYP450), and was verified to catalyze the <i>trans</i>-cinnamic acid to <i>p</i>-coumaric acid transformation by whole-cell bioconversion, HPLC detection and LC-MS analysis as well. Further, with phenylalanine ammonia-lyase 1 of <i>A. thaliana</i>, <i>p</i>-coumaric acid was de novo biosynthesized from glucose as the sole carbon source via the phenylalanine route in the recombinant <i>E. coli</i> cells. By regulating the level of intracellular NADPH, the production of <i>p</i>-coumaric acid was dramatically improved by 9.18-fold, and achieved with a titer of 156.09 μM in shake flasks. The recombinant cells harboring functional <i>Lau</i>C4H afforded a promising chassis for biological production of <i>p</i>-coumaric acid, even other derivatives, via a plant CYP450-involved pathway. |
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| last_indexed | 2024-04-13T00:17:32Z |
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| spelling | doaj.art-038e8e06be3b4648923030bbc5e463f12022-12-22T03:10:53ZengMDPI AGMolecules1420-30492018-12-012312318510.3390/molecules23123185molecules23123185De Novo Biosynthesis of <i>p</i>-Coumaric Acid in <i>E. coli</i> with a <i>trans</i>-Cinnamic Acid 4-Hydroxylase from the Amaryllidaceae Plant <i>Lycoris aurea</i>Yikui Li0Jie Li1Binbin Qian2Li Cheng3Sheng Xu4Ren Wang5Jiangsu Key Laboratory for the Research and Utilization of Plant Resources, Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, ChinaJiangsu Key Laboratory for the Research and Utilization of Plant Resources, Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, ChinaJiangsu Key Laboratory for the Research and Utilization of Plant Resources, Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, ChinaJiangsu Key Laboratory for the Research and Utilization of Plant Resources, Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, ChinaJiangsu Key Laboratory for the Research and Utilization of Plant Resources, Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, ChinaJiangsu Key Laboratory for the Research and Utilization of Plant Resources, Institute of Botany, Jiangsu Province and Chinese Academy of Sciences, Nanjing 210014, China<i>p</i>-Coumaric acid is a commercially available phenolcarboxylic acid with a great number of important applications in the nutraceutical, pharmaceutical, material and chemical industries. <i>p</i>-Coumaric acid has been biosynthesized in some engineered microbes, but the potential of the plant CYP450-involved biosynthetic route has not investigated in <i>Escherichia coli</i>. In the present study, a novel <i>trans</i>-cinnamic acid 4-hydroxylase (C4H) encoding the <i>Lau</i>C4H gene was isolated from <i>Lycoris aurea</i> (L’ Hér.) Herb via rapid amplification of cDNA ends. Then, <i>N</i>-terminal 28 amino acids of <i>Lau</i>C4H were characterized, for the subcellular localization, at the endoplasmic reticulum membrane in protoplasts of <i>Arabidopsis thaliana</i>. In <i>E. coli</i>, <i>Lau</i>C4H without the <i>N</i>-terminal membrane anchor region was functionally expressed when fused with the redox partner of <i>A. thaliana</i> cytochrome P450 enzyme (CYP450), and was verified to catalyze the <i>trans</i>-cinnamic acid to <i>p</i>-coumaric acid transformation by whole-cell bioconversion, HPLC detection and LC-MS analysis as well. Further, with phenylalanine ammonia-lyase 1 of <i>A. thaliana</i>, <i>p</i>-coumaric acid was de novo biosynthesized from glucose as the sole carbon source via the phenylalanine route in the recombinant <i>E. coli</i> cells. By regulating the level of intracellular NADPH, the production of <i>p</i>-coumaric acid was dramatically improved by 9.18-fold, and achieved with a titer of 156.09 μM in shake flasks. The recombinant cells harboring functional <i>Lau</i>C4H afforded a promising chassis for biological production of <i>p</i>-coumaric acid, even other derivatives, via a plant CYP450-involved pathway.https://www.mdpi.com/1420-3049/23/12/3185<i>p</i>-coumaric acid<i>trans</i>-cinnamic acid 4-hydroxylase<i>Lycoris aurea</i><i>Escherichia coli</i>synthetic biology |
| spellingShingle | Yikui Li Jie Li Binbin Qian Li Cheng Sheng Xu Ren Wang De Novo Biosynthesis of <i>p</i>-Coumaric Acid in <i>E. coli</i> with a <i>trans</i>-Cinnamic Acid 4-Hydroxylase from the Amaryllidaceae Plant <i>Lycoris aurea</i> Molecules <i>p</i>-coumaric acid <i>trans</i>-cinnamic acid 4-hydroxylase <i>Lycoris aurea</i> <i>Escherichia coli</i> synthetic biology |
| title | De Novo Biosynthesis of <i>p</i>-Coumaric Acid in <i>E. coli</i> with a <i>trans</i>-Cinnamic Acid 4-Hydroxylase from the Amaryllidaceae Plant <i>Lycoris aurea</i> |
| title_full | De Novo Biosynthesis of <i>p</i>-Coumaric Acid in <i>E. coli</i> with a <i>trans</i>-Cinnamic Acid 4-Hydroxylase from the Amaryllidaceae Plant <i>Lycoris aurea</i> |
| title_fullStr | De Novo Biosynthesis of <i>p</i>-Coumaric Acid in <i>E. coli</i> with a <i>trans</i>-Cinnamic Acid 4-Hydroxylase from the Amaryllidaceae Plant <i>Lycoris aurea</i> |
| title_full_unstemmed | De Novo Biosynthesis of <i>p</i>-Coumaric Acid in <i>E. coli</i> with a <i>trans</i>-Cinnamic Acid 4-Hydroxylase from the Amaryllidaceae Plant <i>Lycoris aurea</i> |
| title_short | De Novo Biosynthesis of <i>p</i>-Coumaric Acid in <i>E. coli</i> with a <i>trans</i>-Cinnamic Acid 4-Hydroxylase from the Amaryllidaceae Plant <i>Lycoris aurea</i> |
| title_sort | de novo biosynthesis of i p i coumaric acid in i e coli i with a i trans i cinnamic acid 4 hydroxylase from the amaryllidaceae plant i lycoris aurea i |
| topic | <i>p</i>-coumaric acid <i>trans</i>-cinnamic acid 4-hydroxylase <i>Lycoris aurea</i> <i>Escherichia coli</i> synthetic biology |
| url | https://www.mdpi.com/1420-3049/23/12/3185 |
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