Cloning and characterization of 2-C-methyl-D-erythritol 2,4-cyclodiphosphate synthase (MECS) gene from Ginkgo biloba

Ginkgo biloba contains secondary metabolites with interesting pharmacological properties, including highly modified diterpenoid ginkgolide, potent and selective antagonist of platelet-activating factor. 2-C-Methyl-D-erythritol 2,4-cyclodiphosphate synthase gene (GbMECS) involved in ginkgolide biosynthesis pathway was cloned and characterized from G. biloba embryonic roots, and the full open reading frame was deduced as protein consisting of 238 amino acid residues. Putative mature protein with a 179 residue-long sequence, obtained by deleting N-terminal chloroplast transit peptide region composed of 59 amino acid residues, rescued Esherichia coli NMW26, an E. coli knock-out mutant of ygbB (EcMECS). Transcription levels of GbMECS were two-fold higher in embryo roots compared to leaves. When full-length GbMECS with chloroplast transit peptide sequence was fused to green fluorescent protein gene (GFP), and transiently expressed in Arabidopsis thaliana protoplast, green fluorescence was found in chloroplast, indication of protein transportation into plastid.