Molecular cloning and functional identification of a cDNA encoding 4-hydroxy-3-methylbut-2-enyl diphosphate reductase from Tripterygium wilfordii
The 4-hydroxy-3-methylbut-2-enyl diphosphate reductase (HDR) is the last step key enzyme of the methylerythritol phosphate (MEP) pathway, synthesizing isopentenyl diphosphate and its allyl isomer dimethylallyl diphosphate, which is important for regulation of isoprenoid biosynthesis. Here the full-l...
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Elsevier
2017-03-01
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| Series: | Acta Pharmaceutica Sinica B |
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| Online Access: | http://www.sciencedirect.com/science/article/pii/S2211383516303136 |
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| author | Qiqing Cheng Yuru Tong Zihao Wang Ping Su Wei Gao Luqi Huang |
| author_facet | Qiqing Cheng Yuru Tong Zihao Wang Ping Su Wei Gao Luqi Huang |
| author_sort | Qiqing Cheng |
| collection | DOAJ |
| description | The 4-hydroxy-3-methylbut-2-enyl diphosphate reductase (HDR) is the last step key enzyme of the methylerythritol phosphate (MEP) pathway, synthesizing isopentenyl diphosphate and its allyl isomer dimethylallyl diphosphate, which is important for regulation of isoprenoid biosynthesis. Here the full-length cDNA of HDR, designated TwHDR (GenBank Accession No. KJ933412.1), was isolated from Tripterygium wilfordii for the first time. TwHDR has an open reading frame (ORF) of 1386 bp encoding 461 amino acids. TwHDR exhibits high homology with HDRs of other plants, with an N-terminal conserved domain and three conserved cysteine residues. TwHDR cDNA was cloned into an expression vector and transformed into an Escherichia coli hdr mutant. Since loss-of-function E.coli hdr mutant is lethal, the result showed that transformation of TwHDR cDNA rescued the E.coli hdr mutant. This complementation assay suggests that the TwHDR cDNA encodes a functional HDR enzyme. The expression of TwHDR was induced by methyl-jasmonate (MJ) in T. wilfordii suspension cells. The expression of TwHDR reached the highest level after 1 h of MJ treatment. These results indicate that we have identified a functional TwHDR enzyme, which may play a pivotal role in the biosynthesis of diterpenoid triptolide in T. wilfordii. |
| first_indexed | 2024-12-11T08:46:03Z |
| format | Article |
| id | doaj.art-f38864ab7156411aa850d4caa97fe4b9 |
| institution | Directory Open Access Journal |
| issn | 2211-3835 2211-3843 |
| language | English |
| last_indexed | 2024-12-11T08:46:03Z |
| publishDate | 2017-03-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Acta Pharmaceutica Sinica B |
| spelling | doaj.art-f38864ab7156411aa850d4caa97fe4b92022-12-22T01:14:08ZengElsevierActa Pharmaceutica Sinica B2211-38352211-38432017-03-017220821410.1016/j.apsb.2016.12.002Molecular cloning and functional identification of a cDNA encoding 4-hydroxy-3-methylbut-2-enyl diphosphate reductase from Tripterygium wilfordiiQiqing Cheng0Yuru Tong1Zihao Wang2Ping Su3Wei Gao4Luqi Huang5School of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, ChinaSchool of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, ChinaSchool of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, ChinaSchool of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, ChinaSchool of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, ChinaNational Resource Center for Chinese Materia Medica, Academy of Chinese Medical Sciences, Beijing 100700, ChinaThe 4-hydroxy-3-methylbut-2-enyl diphosphate reductase (HDR) is the last step key enzyme of the methylerythritol phosphate (MEP) pathway, synthesizing isopentenyl diphosphate and its allyl isomer dimethylallyl diphosphate, which is important for regulation of isoprenoid biosynthesis. Here the full-length cDNA of HDR, designated TwHDR (GenBank Accession No. KJ933412.1), was isolated from Tripterygium wilfordii for the first time. TwHDR has an open reading frame (ORF) of 1386 bp encoding 461 amino acids. TwHDR exhibits high homology with HDRs of other plants, with an N-terminal conserved domain and three conserved cysteine residues. TwHDR cDNA was cloned into an expression vector and transformed into an Escherichia coli hdr mutant. Since loss-of-function E.coli hdr mutant is lethal, the result showed that transformation of TwHDR cDNA rescued the E.coli hdr mutant. This complementation assay suggests that the TwHDR cDNA encodes a functional HDR enzyme. The expression of TwHDR was induced by methyl-jasmonate (MJ) in T. wilfordii suspension cells. The expression of TwHDR reached the highest level after 1 h of MJ treatment. These results indicate that we have identified a functional TwHDR enzyme, which may play a pivotal role in the biosynthesis of diterpenoid triptolide in T. wilfordii.http://www.sciencedirect.com/science/article/pii/S2211383516303136Tripterygium wilfordiiTriptolide4-Hydroxy-3-methylbut-2-enyl diphosphate reductaseComplementationGene expression |
| spellingShingle | Qiqing Cheng Yuru Tong Zihao Wang Ping Su Wei Gao Luqi Huang Molecular cloning and functional identification of a cDNA encoding 4-hydroxy-3-methylbut-2-enyl diphosphate reductase from Tripterygium wilfordii Acta Pharmaceutica Sinica B Tripterygium wilfordii Triptolide 4-Hydroxy-3-methylbut-2-enyl diphosphate reductase Complementation Gene expression |
| title | Molecular cloning and functional identification of a cDNA encoding 4-hydroxy-3-methylbut-2-enyl diphosphate reductase from Tripterygium wilfordii |
| title_full | Molecular cloning and functional identification of a cDNA encoding 4-hydroxy-3-methylbut-2-enyl diphosphate reductase from Tripterygium wilfordii |
| title_fullStr | Molecular cloning and functional identification of a cDNA encoding 4-hydroxy-3-methylbut-2-enyl diphosphate reductase from Tripterygium wilfordii |
| title_full_unstemmed | Molecular cloning and functional identification of a cDNA encoding 4-hydroxy-3-methylbut-2-enyl diphosphate reductase from Tripterygium wilfordii |
| title_short | Molecular cloning and functional identification of a cDNA encoding 4-hydroxy-3-methylbut-2-enyl diphosphate reductase from Tripterygium wilfordii |
| title_sort | molecular cloning and functional identification of a cdna encoding 4 hydroxy 3 methylbut 2 enyl diphosphate reductase from tripterygium wilfordii |
| topic | Tripterygium wilfordii Triptolide 4-Hydroxy-3-methylbut-2-enyl diphosphate reductase Complementation Gene expression |
| url | http://www.sciencedirect.com/science/article/pii/S2211383516303136 |
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