Molecular cloning and characterization of O-methyltransferases from the flower buds of Iris hollandica
| Autor: | Yoshikazu Tanaka, Hiroaki Okuhara, Noriko Yoshihara, Tsutomu Yabuya, Masako Fukuchi-Mizutani |
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| Rok vydání: | 2008 |
| Předmět: |
Iris × hollandica
Iris Plant Physiology Molecular Sequence Data Flowers Plant Science Molecular cloning Biology Gene Expression Regulation Enzymologic chemistry.chemical_compound Gene Expression Regulation Plant Complementary DNA Caffeic acid Amino Acid Sequence RNA Messenger Cloning Molecular Peptide sequence Chromatography High Pressure Liquid Phylogeny chemistry.chemical_classification Sequence Homology Amino Acid Phenylpropanoid Reverse Transcriptase Polymerase Chain Reaction cDNA library Temperature food and beverages Hydrogen-Ion Concentration Protein O-Methyltransferase S-Adenosylhomocysteine Molecular biology Recombinant Proteins Amino acid chemistry Biochemistry Biocatalysis Electrophoresis Polyacrylamide Gel Sequence Alignment Agronomy and Crop Science |
| Zdroj: | Journal of Plant Physiology. 165:415-422 |
| ISSN: | 0176-1617 |
| DOI: | 10.1016/j.jplph.2006.12.002 |
| Popis: | In plants, O-methyltransferases (OMTs) play an important role in methylation of secondary metabolites, especially flavonoids and other phenylpropanoids, and two cDNA clones, IhOMT1 and IhOMT2 (Iris hollandica OMT), encoding OMTs were successfully isolated from a cDNA library of flower buds of I. hollandica. IhOMT1 encodes an open reading frame (ORF) of 365 amino acids with calculated molecular mass of 40,193Da and isoelectric point (pI) of 5.54, while IhOMT2, which shares 31.5% amino acid sequence identity with IhOMT1, encodes 369 amino acids with calculated molecular mass of 40,385Da and pI of 5.50. In addition, the molecular masses of both recombinant IhOMT1 and IhOMT2 proteins were estimated to be about 40kDa by protein gel blot analysis. Characterization of the enzymatic properties using the recombinant IhOMT1 protein confirmed that IhOMT1 cDNA encodes a S-adenosyl-L-methionine (SAM)-dependent caffeic acid 3-OMT, which catalyzes the transfer of the methyl moiety from SAM to caffeic acid to form ferulic acid. Its optimum activity was observed at pH 7.5-8.0 and at 35 degrees C. This is the first report of the isolation and characterization of a COMT cDNA clone involved in the phenylpropanoid biosynthesis of Iridaceae plants. In contrast, IhOMT2 showed no activity in SAM-dependent assays for various phenylpropanoids. |
| Databáze: | OpenAIRE |
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