O-methylation of flavonoid substrates by a partially purified enzyme from soybean cell suspension cultures

Abstract

A methyltransferase, which catalyzes the methylation of luteolin ( K m , 16 μM) using S-adenosyl- l-methionine as the methyl donor, has been purified about 38-fold from cell suspension cultures of soybean ( Glycine max L., var. Mandarin). The following 3,4-dihydroxy phenolic compounds were also methylated: luteolin 7- O-glucoside ( K m , 28 μ m), quercetin ( K m , 35 μ m), eriodictyol ( K m , 75 μ m), 5-hydroxyferulic acid ( K m , 227 μ m), dihydroquercetin ( K m , 435 μ m), and caffeic acid ( K m , 770 μ m). Rutin and quercetin 3- O-glucoside were poor substrates. Methylation proceeded only in the meta position. The enzyme was unable to catalyze the methylation of p-coumaric acid, m-coumaric acid, ferulic acid, isoferulic acid, sinapic acid, apigenin, or naringenin. While the isoflavones biochanin A and daidzein did not serve as substrates, texasin (6,7-dihydroxy-3′-methoxyisoflavone) was methylated ( K m , 35 μ m). The methylation of caffeic acid and quercetin showed a pH optimum of 8.6–8.9. The enzyme required Mg 2+ ions for maximum activity (approximately 1 m m) and could be totally inhibited by EDTA (10 m m). The K m for S-adenosyl- l-methionine was 11 μ m. S-Adenosyl- l-homocysteine inhibited the methylation of luteolin by S-adenosyl- l-methionine.