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KMID : 0380220060390050502
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Journal of Biochemistry and Molecular Biology
2006 Volume.39 No. 5 p.502 ~ p.510
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Molecular Cloning, Characterization and Functional Analysis of a 2C-methyl-D-erythritol 2, 4-cyclodiphosphate Synthase Gene from Ginkgo biloba
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Gao Shi
Lin Juan
Liu Xuefen
Deng Zhongxiang
Li Yingjun
Sun Xiaofen
Tang Kexuan
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Abstract
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2C-methyl-D-erythritol 2, 4-cyclodiphosphate synthase (MECPS, EC: 4.6.1.12) is the fifth enzyme of the nonmevalonate terpenoid pathway for isopentenyl diphosphate biosynthesis and is involved in the methylerythritol phosphate (MEP) pathway for ginkgolide biosynthesis. The full-length mecps cDNA sequence (designated as Gbmecps) was cloned and characterized for the first time from gymnosperm plant species, Ginkgo biloba, using RACE (rapid amplification of cDNA ends) technique. The full-length cDNA of Gbmecps was 874 bp containing a 720 bp open reading frame (ORF) encoding a peptide of 239 amino acids with a calculated molecular mass of 26.03 kDa and an isoelectric point of 8.83. Comparative and bioinformatic analyses revealed that GbMECPS showed extensive homology with MECPSs from other species and contained conserved residues owned by the MECPS protein family.Phylogenetic analysis indicated that GbMECPS was more ancient than other plant MECPSs. Tissue expression pattern analysis indicated that GbMECPS expressed the highest in roots, followed by in leaves, and the lowest in seeds. The color complementation assay indicated that GbMECPS could accelerate the accumulation of &bgr;-carotene. The cloning, characterization and functional analysis of GbMECPS will be helpful to understand more about the role of MECPS involved in the ginkgolides biosynthesis at the molecular level.
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KEYWORD
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Color complementation assay, 2C-methyl-D-erythritol 2, 4-cyclodiphosphate synthase (MECPS), Ginkgo biloba, Ginkgolides biosynthesis, RACE
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