Molecular cloning, characterization and functional analysis of a 2C-methyl- D-erythritol 2, 4-cyclodiphosphate synthase gene from ginkgo biloba.
2C-methyl-D-erythritol 2, 4-cyclodiphosphate synthase (MECPS, EC: 4.6.1.12) is the fifth enzyme of the non-mevalonate 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 beta-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.
Gao
Shi
S
State Key Laboratory of Genetic Engineering, School of Life Sciences, Morgan-Tan International Center for Life Sciences, Fudan-SJTU-Nottingham Plant Biotechnology R&D Center, Fudan University, Shanghai 200433, China.
Lin
Juan
J
Liu
Xuefen
X
Deng
Zhongxiang
Z
Li
Yingjun
Y
Sun
Xiaofen
X
Tang
Kexuan
K
D000595:Amino Acid Sequence; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D019295:Computational Biology; D004896:Erythritol / Q000031:analogs & derivatives* / Q000096:biosynthesis; D019143:Evolution, Molecular; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D020441:Ginkgo biloba / Q000201:enzymology / Q000235:genetics*; D046934:Ginkgolides / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D019761:Phosphorus-Oxygen Lyases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D010802:Phylogeny; D010940:Plant Proteins; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016415:Sequence Alignment
D046934:Ginkgolides; D010940:Plant Proteins; C443879:methyl-D-erythritol 2,4-cyclodiphosphate; D019761:Phosphorus-Oxygen Lyases; D004896:Erythritol