title: Functional characterization of a Mg(2+)-dependent O-methyltransferase with coumarin as preferred substrate from the liverwort Plagiochasma appendiculatum.
abstract: Coumarins (1,2-benzopyrones), which originate via the phenylpropanoid pathway, are found ubiquitously in plants and make an essential contribution to the health of the plant. Some natural coumarins have been used as human therapeutics. However, the details of their biosynthesis are still largely unknown. Scopoletin is derived from either esculetin or feruloyl CoA according to the plant species involved. Here, a gene encoding a O-methyltransferase (PaOMT2) was isolated from the liverwort species Plagiochasma appendiculatum (Aytoniaceae) through transcriptome sequencing. The purified recombinant enzyme catalyzed the methylation of esculetin, generating scopoletin and isoscopoletin. Kinetic analysis shows that the construct from the second Met in PaOMT2 had a catalytic efficiency for esculetin (Kcat/Km) of about half that of the full length PaOMT2, while the Kms of two enzymes were similar. The catalytic capacities of the studied protein suggest that two routes to scopoletin might co-exist in liverworts in that the enzyme involved in the methylation process participates in both paths, but especially the route from esculetin. The transient expression of a PaOMT2-GFP fusion in tobacco demonstrated that PaOMT2 is directed to the cytoplasm.
authors.affiliation: Key Laboratory of Chemical Biology of Natural Products, Ministry of Education, School of Pharmaceutical Sciences, Shandong University, Jinan, 250012, China.
authors.forename: Rui-Xue
authors.initials: RX
authors.lastname: Xu
authors.affiliation: Key Laboratory of Chemical Biology of Natural Products, Ministry of Education, School of Pharmaceutical Sciences, Shandong University, Jinan, 250012, China.
authors.forename: Shuai
authors.initials: S
authors.lastname: Gao
authors.affiliation: Key Laboratory of Chemical Biology of Natural Products, Ministry of Education, School of Pharmaceutical Sciences, Shandong University, Jinan, 250012, China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Zhao
authors.affiliation: Key Laboratory of Chemical Biology of Natural Products, Ministry of Education, School of Pharmaceutical Sciences, Shandong University, Jinan, 250012, China.
authors.forename: Hong-Xiang
authors.initials: HX
authors.lastname: Lou
authors.affiliation: Key Laboratory of Chemical Biology of Natural Products, Ministry of Education, School of Pharmaceutical Sciences, Shandong University, Jinan, 250012, China. Electronic address: aixiacheng@sdu.edu.cn.
authors.forename: Ai-Xia
authors.initials: AX
authors.lastname: Cheng
mesh_terms: D000595:Amino Acid Sequence; D053898:Biosynthetic Pathways / Q000187:drug effects; D003001:Cloning, Molecular; D003374:Coumarins / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D035926:Hepatophyta / Q000187:drug effects / Q000201:enzymology*; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology*; D008780:Methyltransferases / Q000737:chemistry / Q000378:metabolism*; D010802:Phylogeny; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D012603:Scopoletin / Q000737:chemistry / Q000378:metabolism; D016415:Sequence Alignment; D013347:Subcellular Fractions / Q000378:metabolism; D013379:Substrate Specificity / Q000187:drug effects; D014468:Umbelliferones / Q000378:metabolism
chemical_list: D003374:Coumarins; D011994:Recombinant Proteins; D014468:Umbelliferones; C030123:coumarin; D008780:Methyltransferases; D008274:Magnesium; D012603:Scopoletin; C007628:esculetin
keywords: Coumarins; Esculetin; Liverworts; O-methyltransferase; Plagiochasma appendiculatum; Scopoletin