title: cDNA cloning and characterization of a 3'/5'-O-methyltransferase for partially methylated flavonols from Chrysosplenium americanum.
abstract: Enzymatic O-methylation of plant secondary metabolites is an important mechanism for the inactivation of reactive hydroxyl groups and for the modification of their solubility. A cDNA clone (pFOMT3') encoding the gene for the 3'/5'-O-methylation of partially methylated flavonols was isolated from Chrysosplenium americanum (Saxifragaceae). We used a PCR fragment obtained with degenerate oligonucleotides designed from conserved regions of various O-methyltransferases (OMTs). The pFOMT3' cDNA sequence shows about 67-85% similarity to other plant OMT sequences. The recombinant protein expresses strict specificity for positions 3'/5' (meta) of partially methylated flavonols, but does not accept quercetin or caffeic acid for further methylation. Southern blot analysis of the genomic DNA probed with an OMT sequence suggests the presence of a number of related genes in this species, consistent with the multiple enzymatic methylations involved in the biosynthesis of polymethylated flavonols in this plant.
authors.affiliation: Plant Biochemistry Laboratory, Department of Biology, Concordia University, Montréal, Québec, Canada.
authors.forename: A
authors.initials: A
authors.lastname: Gauthier
authors.forename: P J
authors.initials: PJ
authors.lastname: Gulick
authors.forename: R K
authors.initials: RK
authors.lastname: Ibrahim
mesh_terms: D000595:Amino Acid Sequence; D015139:Blotting, Southern; D002109:Caffeic Acids / Q000378:metabolism; D002394:Catechol O-Methyltransferase / Q000235:genetics; D003001:Cloning, Molecular; D017931:DNA Primers; D018076:DNA, Complementary / Q000235:genetics; D005419:Flavonoids / Q000378:metabolism*; D044948:Flavonols; D015723:Gene Library; D017343:Genes, Plant; D008745:Methylation; D008780:Methyltransferases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010940:Plant Proteins; D010944:Plants / Q000201:enzymology* / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011794:Quercetin / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D013379:Substrate Specificity
chemical_list: D002109:Caffeic Acids; D017931:DNA Primers; D018076:DNA, Complementary; D005419:Flavonoids; D044948:Flavonols; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; D011794:Quercetin; D008780:Methyltransferases; C114844:trimethylflavonol 3'-O-methyltransferase, Chrysosplenium americanum; D002394:Catechol O-Methyltransferase; C040048:caffeic acid