Cloning, characterization, and expression of 1-deoxy-D-xylulose-5-phosphate reductoisomerase gene from Dioscorea zingiberensis

Abstract

Diosgenin, mainly extracted from some Dioscorea species, is the most important starting material for the production of steroidal drugs. It is believed that diosgenin in Dioscorea is synthesized from isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP, the isomer of IPP) produced by the cytosolic mevalonate pathway. So far, the possibility of the plastidic 2-C-methyl-D-erythritol 4-phosphate (MEP) pathway for biosynthesis of diosgenin remains unclear. The key enzyme in the MEP pathway is 1-deoxy-D-xylulose 5-phosphate-reductoisomerase (DXR, EC: 1.1.1.267). In this study, a DXR gene, named DzDXR (GenBank accession number KY131955), was isolated from Dioscorea zingiberensis. The DzDXR has an open reading frame of 1 413 bp encoding a protein of 470 amino acid residues. The function of DzDXR was verified by a colour enhancement assay in the Escherichia coli cells harbouring the plasmid pAC-BETA. Bioinformatic analyses revealed that DzDXR had a putative plastid transit peptide at the N-terminal region and was highly homologous to other plant DXRs, especially to those in monocotyledons. During the growth period of D. zingiberensis, the expression of DzDXR was found significantly high in leaves and very low in tubers, and the highest expression was observed in mature leaves in summer. In contrast to the DzDXR transcription, diosgenin was present predominantly in tubers and in minute quantities in leaves. Because diosgenin is very likely formed mainly in the cytosol of mature leaf cells of Dioscorea and the plastidic IPP and DMAPP produced by the MEP pathway can be transported into the cytosol, the consistently high expression of DzDXR detected in mature leaf of D. zingiberensis implies that the MEP pathway might play a significant role in diosgenin biosynthesis.