Enhancing Tropane Alkaloid Production Based on the Functional Identification of Tropine-Forming Reductase in Scopolia lurida, a Tibetan Medicinal Plant
Scopolia lurida, a native herbal plant species in Tibet, is one of the most effective producers of tropane alkaloids. However, the tropane alkaloid biosynthesis in this plant species of interest has yet to be studied at the molecular, biochemical, and biotechnological level. Here, we report on the i...
| Main Authors: | , , , , , , , , , , |
|---|---|
| Format: | Article |
| Language: | English |
| Published: |
Frontiers Media S.A.
2017-10-01
|
| Series: | Frontiers in Plant Science |
| Subjects: | |
| Online Access: | http://journal.frontiersin.org/article/10.3389/fpls.2017.01745/full |
| _version_ | 1818132231581859840 |
|---|---|
| author | Kaihui Zhao Kaihui Zhao Junlan Zeng Tengfei Zhao Haoxing Zhang Fei Qiu Chunxian Yang Lingjiang Zeng Xiaoqiang Liu Min Chen Xiaozhong Lan Xiaozhong Lan Zhihua Liao Zhihua Liao |
| author_facet | Kaihui Zhao Kaihui Zhao Junlan Zeng Tengfei Zhao Haoxing Zhang Fei Qiu Chunxian Yang Lingjiang Zeng Xiaoqiang Liu Min Chen Xiaozhong Lan Xiaozhong Lan Zhihua Liao Zhihua Liao |
| author_sort | Kaihui Zhao |
| collection | DOAJ |
| description | Scopolia lurida, a native herbal plant species in Tibet, is one of the most effective producers of tropane alkaloids. However, the tropane alkaloid biosynthesis in this plant species of interest has yet to be studied at the molecular, biochemical, and biotechnological level. Here, we report on the isolation and characterization of a putative short chain dehydrogenase (SDR) gene. Sequence analysis showed that SlTRI belonged to the SDR family. Phylogenetic analysis revealed that SlTRI was clustered with the tropine-forming reductases. SlTRI and the other TA-biosynthesis genes, including putrescine N-methyltransferase (SlPMT) and hyoscyamine 6β-hydroxylase (SlH6H), were preferably or exclusively expressed in the S. lurida roots. The tissue profile of SlTRI suggested that this gene might be involved in tropane alkaloid biosynthesis. By using GC-MS, SlTRI was shown to catalyze the tropinone reduction to yield tropine, the key intermediate of tropane alkaloids. With the purified recombinant SlTRI from Escherichiacoli, an enzymatic assay was carried out; its result indicated that SlTRI was a tropine-forming reductase. Finally, the role of SlTRI in promoting the tropane alkaloid biosynthesis was confirmed through metabolic engineering in S. lurida. Specifically, hairy root cultures of S. lurida were established to investigate the effects of SlTRI overexpression on tropane alkaloid accumulation. In the SlTRI-overexpressing root cultures, the hyoscyamine contents were 1.7- to 2.9-fold higher than those in control while their corresponding scopolamine contents were likewise elevated. In summary, this functional identification of SlTRI has provided for a better understanding of tropane alkaloid biosynthesis. It also provides a candidate gene for enhancing tropane alkaloid biosynthesis in S. lurida via metabolic engineering. |
| first_indexed | 2024-12-11T08:33:33Z |
| format | Article |
| id | doaj.art-57fcf9b5c2d548f3b5b467ec456f3027 |
| institution | Directory Open Access Journal |
| issn | 1664-462X |
| language | English |
| last_indexed | 2024-12-11T08:33:33Z |
| publishDate | 2017-10-01 |
| publisher | Frontiers Media S.A. |
| record_format | Article |
| series | Frontiers in Plant Science |
| spelling | doaj.art-57fcf9b5c2d548f3b5b467ec456f30272022-12-22T01:14:23ZengFrontiers Media S.A.Frontiers in Plant Science1664-462X2017-10-01810.3389/fpls.2017.01745266196Enhancing Tropane Alkaloid Production Based on the Functional Identification of Tropine-Forming Reductase in Scopolia lurida, a Tibetan Medicinal PlantKaihui Zhao0Kaihui Zhao1Junlan Zeng2Tengfei Zhao3Haoxing Zhang4Fei Qiu5Chunxian Yang6Lingjiang Zeng7Xiaoqiang Liu8Min Chen9Xiaozhong Lan10Xiaozhong Lan11Zhihua Liao12Zhihua Liao13Key Laboratory of Eco-Environments in Three Gorges Reservoir Region (Ministry of Education), Chongqing Key Laboratory of Plant Ecology and Resources Research in Three Gorges Reservoir Region, SWU-TAAHC Medicinal Plant Joint R&D Centre, School of Life Sciences, Southwest University, Chongqing, ChinaTAAHC-SWU Medicinal Plant Joint R&D Centre, Tibetan Collaborative Innovation Centre of Agricultural and Animal Husbandry Resources, Tibet Agricultural and Animal Husbandry College, Nyingchi, ChinaKey Laboratory of Eco-Environments in Three Gorges Reservoir Region (Ministry of Education), Chongqing Key Laboratory of Plant Ecology and Resources Research in Three Gorges Reservoir Region, SWU-TAAHC Medicinal Plant Joint R&D Centre, School of Life Sciences, Southwest University, Chongqing, ChinaKey Laboratory of Eco-Environments in Three Gorges Reservoir Region (Ministry of Education), Chongqing Key Laboratory of Plant Ecology and Resources Research in Three Gorges Reservoir Region, SWU-TAAHC Medicinal Plant Joint R&D Centre, School of Life Sciences, Southwest University, Chongqing, ChinaCollege of Life Sciences and Oceanography, Shenzhen University, Shenzhen, ChinaKey Laboratory of Eco-Environments in Three Gorges Reservoir Region (Ministry of Education), Chongqing Key Laboratory of Plant Ecology and Resources Research in Three Gorges Reservoir Region, SWU-TAAHC Medicinal Plant Joint R&D Centre, School of Life Sciences, Southwest University, Chongqing, ChinaKey Laboratory of Eco-Environments in Three Gorges Reservoir Region (Ministry of Education), Chongqing Key Laboratory of Plant Ecology and Resources Research in Three Gorges Reservoir Region, SWU-TAAHC Medicinal Plant Joint R&D Centre, School of Life Sciences, Southwest University, Chongqing, ChinaKey Laboratory of Eco-Environments in Three Gorges Reservoir Region (Ministry of Education), Chongqing Key Laboratory of Plant Ecology and Resources Research in Three Gorges Reservoir Region, SWU-TAAHC Medicinal Plant Joint R&D Centre, School of Life Sciences, Southwest University, Chongqing, ChinaKey Laboratory of Eco-Environments in Three Gorges Reservoir Region (Ministry of Education), Chongqing Key Laboratory of Plant Ecology and Resources Research in Three Gorges Reservoir Region, SWU-TAAHC Medicinal Plant Joint R&D Centre, School of Life Sciences, Southwest University, Chongqing, ChinaSWU-TAAHC Medicinal Plant Joint R&D Centre, College of Pharmaceutical Sciences, Southwest University, Chongqing, ChinaTAAHC-SWU Medicinal Plant Joint R&D Centre, Tibetan Collaborative Innovation Centre of Agricultural and Animal Husbandry Resources, Tibet Agricultural and Animal Husbandry College, Nyingchi, ChinaState Key Laboratory Breeding Base of Dao-di Herbs, National Resource Centre for Chinese Materia Medica, China Academy of Chinese Medical Sciences, Beijing, ChinaKey Laboratory of Eco-Environments in Three Gorges Reservoir Region (Ministry of Education), Chongqing Key Laboratory of Plant Ecology and Resources Research in Three Gorges Reservoir Region, SWU-TAAHC Medicinal Plant Joint R&D Centre, School of Life Sciences, Southwest University, Chongqing, ChinaTAAHC-SWU Medicinal Plant Joint R&D Centre, Tibetan Collaborative Innovation Centre of Agricultural and Animal Husbandry Resources, Tibet Agricultural and Animal Husbandry College, Nyingchi, ChinaScopolia lurida, a native herbal plant species in Tibet, is one of the most effective producers of tropane alkaloids. However, the tropane alkaloid biosynthesis in this plant species of interest has yet to be studied at the molecular, biochemical, and biotechnological level. Here, we report on the isolation and characterization of a putative short chain dehydrogenase (SDR) gene. Sequence analysis showed that SlTRI belonged to the SDR family. Phylogenetic analysis revealed that SlTRI was clustered with the tropine-forming reductases. SlTRI and the other TA-biosynthesis genes, including putrescine N-methyltransferase (SlPMT) and hyoscyamine 6β-hydroxylase (SlH6H), were preferably or exclusively expressed in the S. lurida roots. The tissue profile of SlTRI suggested that this gene might be involved in tropane alkaloid biosynthesis. By using GC-MS, SlTRI was shown to catalyze the tropinone reduction to yield tropine, the key intermediate of tropane alkaloids. With the purified recombinant SlTRI from Escherichiacoli, an enzymatic assay was carried out; its result indicated that SlTRI was a tropine-forming reductase. Finally, the role of SlTRI in promoting the tropane alkaloid biosynthesis was confirmed through metabolic engineering in S. lurida. Specifically, hairy root cultures of S. lurida were established to investigate the effects of SlTRI overexpression on tropane alkaloid accumulation. In the SlTRI-overexpressing root cultures, the hyoscyamine contents were 1.7- to 2.9-fold higher than those in control while their corresponding scopolamine contents were likewise elevated. In summary, this functional identification of SlTRI has provided for a better understanding of tropane alkaloid biosynthesis. It also provides a candidate gene for enhancing tropane alkaloid biosynthesis in S. lurida via metabolic engineering.http://journal.frontiersin.org/article/10.3389/fpls.2017.01745/fullenzymatic assaymetabolic engineeringScopolia luridatropane alkaloidstropine-forming reductase |
| spellingShingle | Kaihui Zhao Kaihui Zhao Junlan Zeng Tengfei Zhao Haoxing Zhang Fei Qiu Chunxian Yang Lingjiang Zeng Xiaoqiang Liu Min Chen Xiaozhong Lan Xiaozhong Lan Zhihua Liao Zhihua Liao Enhancing Tropane Alkaloid Production Based on the Functional Identification of Tropine-Forming Reductase in Scopolia lurida, a Tibetan Medicinal Plant Frontiers in Plant Science enzymatic assay metabolic engineering Scopolia lurida tropane alkaloids tropine-forming reductase |
| title | Enhancing Tropane Alkaloid Production Based on the Functional Identification of Tropine-Forming Reductase in Scopolia lurida, a Tibetan Medicinal Plant |
| title_full | Enhancing Tropane Alkaloid Production Based on the Functional Identification of Tropine-Forming Reductase in Scopolia lurida, a Tibetan Medicinal Plant |
| title_fullStr | Enhancing Tropane Alkaloid Production Based on the Functional Identification of Tropine-Forming Reductase in Scopolia lurida, a Tibetan Medicinal Plant |
| title_full_unstemmed | Enhancing Tropane Alkaloid Production Based on the Functional Identification of Tropine-Forming Reductase in Scopolia lurida, a Tibetan Medicinal Plant |
| title_short | Enhancing Tropane Alkaloid Production Based on the Functional Identification of Tropine-Forming Reductase in Scopolia lurida, a Tibetan Medicinal Plant |
| title_sort | enhancing tropane alkaloid production based on the functional identification of tropine forming reductase in scopolia lurida a tibetan medicinal plant |
| topic | enzymatic assay metabolic engineering Scopolia lurida tropane alkaloids tropine-forming reductase |
| url | http://journal.frontiersin.org/article/10.3389/fpls.2017.01745/full |
| work_keys_str_mv | AT kaihuizhao enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant AT kaihuizhao enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant AT junlanzeng enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant AT tengfeizhao enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant AT haoxingzhang enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant AT feiqiu enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant AT chunxianyang enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant AT lingjiangzeng enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant AT xiaoqiangliu enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant AT minchen enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant AT xiaozhonglan enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant AT xiaozhonglan enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant AT zhihualiao enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant AT zhihualiao enhancingtropanealkaloidproductionbasedonthefunctionalidentificationoftropineformingreductaseinscopolialuridaatibetanmedicinalplant |