title: Molecular cloning and functional identification of a cDNA encoding 4-hydroxy-3-methylbut-2-enyl diphosphate reductase fromTripterygium wilfordii.
abstract: 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 ofHDR, designatedTwHDR(GenBank Accession No. KJ933412.1), was isolated fromTripterygium wilfordiifor 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.TwHDRcDNA was cloned into an expression vector and transformed into anEscherichia coli hdrmutant. Since loss-of-functionE.coli hdrmutant is lethal, the result showed that transformation ofTwHDRcDNA rescued theE.coli hdrmutant. This complementation assay suggests that theTwHDRcDNA encodes a functional HDR enzyme. The expression ofTwHDRwas induced by methyl-jasmonate (MJ) inT. wilfordiisuspension cells. The expression ofTwHDRreached 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 inT. wilfordii.
authors.affiliation: School of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, China; National Resource Center for Chinese Materia Medica, Academy of Chinese Medical Sciences, Beijing 100700, China; State Key Laboratory of Quality Research in Chinese Medicines, Macau University of Science and Technology, Macau, China.
authors.forename: Qiqing
authors.initials: Q
authors.lastname: Cheng
authors.affiliation: School of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, China; National Resource Center for Chinese Materia Medica, Academy of Chinese Medical Sciences, Beijing 100700, China.
authors.forename: Yuru
authors.initials: Y
authors.lastname: Tong
authors.affiliation: School of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, China.
authors.forename: Zihao
authors.initials: Z
authors.lastname: Wang
authors.affiliation: School of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, China; National Resource Center for Chinese Materia Medica, Academy of Chinese Medical Sciences, Beijing 100700, China.
authors.forename: Ping
authors.initials: P
authors.lastname: Su
authors.affiliation: School of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Gao
authors.affiliation: National Resource Center for Chinese Materia Medica, Academy of Chinese Medical Sciences, Beijing 100700, China.
authors.forename: Luqi
authors.initials: L
authors.lastname: Huang
keywords: 4-Hydroxy-3-methylbut-2-enyl diphosphate reductase; Complementation; Gene expression; Tripterygium wilfordii; Triptolide