JOURNAL OF SHANDONG UNIVERSITY (HEALTH SCIENCES) ›› 2016, Vol. 54 ›› Issue (12): 20-26.doi: 10.6040/j.issn.1671-7554.0.2016.1137

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Cloning and functional characterization of an O-methyltransferase gene from Conocephalum conicum

LI Dandan, LIU Hui, CHENG Aixia   

  1. Key Laboratory of Chemical Biology of Natural Products, Ministry of Education;
    School of Pharmaceutical Sciences, Shandong University, Jinan 250012, Shandong, China
  • Received:2016-09-09 Online:2016-12-10 Published:2016-12-10

Abstract: Objective To isolate and characterize an O-methyltransferase(OMT)gene from Conocephalum conicum. Methods The full length cDNA sequence of CcOMT1 was obtained by RT-PCR and inserted into the prokaryotic expression plasmid pET32a, and then was transformed into Escherichia coli BL21(DE3)for expression. The recombinant protein was collected by Ni-NTA affinity column and purified for enzyme characterization. The analyses of multiple alignment and phylogenetic tree were performed using DNAMAN 7.0 and MEGA 5.1 softwares. Results Sequence analysis showed that the full length cDNA of CcOMT1 was 837 bp and encoded a 278-aa protein with a calculated molecular weight of 30.95 kDa and an isoelectric point of 5.59. The amino acid sequence of CcOMT1 showed 73.18%, 53.60%, and 44.60% identity with O-methyltransferase(OMT)from Plagiochasma appendiculatum, Medicago sativa and Mesembryanthemum crystallinum, respectively. The purified recombinant protein was able to methylate a variety of flavonoids and phenylpropanoids with aromatic vicinal dihydroxyl groups, and the preferred substrate was quercetin. Conclusion An OMT gene from Conocephalum conicum is cloned and characterized. The present investigation would provide a candidate gene for synthesis of methylated compounds at specific position through enzymatic method.

Key words: Conocephalum conicum, O-methyltransferase, Flavonoids, Enzyme activity assay, Gene cloning

CLC Number: 

  • Q789
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