Substrate preferences of O-methyltransferases in alfalfa suggest new pathways for 3-O-methylation of monolignols

• Published in: The Plant journal : for cell and molecular biology 2001, Vol.25 (2), p.193-202
• Main Authors: PARVATHI, Kota, FANG CHEN, DIANJING GUO, BLOUNT, Jack W, DIXON, Richard A
• Format: Article
• Language: English
• Subjects: Agronomy. Soil science and plant productions; Biological and medical sciences; Catechol O-Methyltransferase - metabolism; Economic plant physiology; Electrophoresis, Polyacrylamide Gel; Enzymes; Fundamental and applied biological sciences. Psychology; Kinetics; Lignin - metabolism; Medicago sativa - enzymology; Metabolism; Methylation; Methyltransferases - metabolism; Nitrogen metabolism and other ones (excepting carbon metabolism); Nutrition. Photosynthesis. Respiration. Metabolism; Plant physiology and development; Plants, Genetically Modified - enzymology; Substrate Specificity
• ISSN: 0960-7412; 1365-313X
• DOI: 10.1046/j.1365-313x.2001.00956.x

Measurement of relative O-methyltransferase activities against all potential substrates in the monolignol pathway in developing alfalfa stem extracts revealed activities in the order: caffeoyl CoA > caffeoyl alcohol > 5-hydroxyferulic acid > caffeoyl aldehyde > 5-hydroxyconiferyl alcohol > 5-hydroxyferuloyl CoA > 5-hydroxyconiferaldehyde > caffeic acid. Maxima for all activities occurred in the seventh internode. In stem extracts from transgenic alfalfa with antisense downregulated caffeoyl CoA O-methyltransferase (CCoAOMT), activities with all substrates except for the two coenzyme A esters were unaffected. In contrast, downregulation of caffeic acid O-methyltransferase (COMT) reduced activities against the non-esterifed substrates in the order: 5-hydroxyconiferyl alcohol > 5-hydroxyferulic acid and caffeoyl alcohol > caffeoyl aldehyde > caffeic acid > 5-hydroxyconiferaldehyde. Recombinant COMT expressed in Escherichia coli exhibited the highest V(max)/K(m) values with 5-hydroxyconiferaldehyde and caffeoyl aldehyde, and the lowest with caffeic acid. These results indicate that COMT is unlikely to methylate caffeic acid during lignin biosynthesis in vivo, and provide enzymatic evidence for an alternative pathway to monolignols involving methylation of caffeoyl aldehyde and/or caffeoyl alcohol by COMT. The concept of independent pathways to guaiacyl and syringyl monolignols is discussed.