Enzymic synthesis of lignin precursors: Purification and properties of the S-adenosyl- l-methionine: Caffeic acid 3- O-methyltransferase from soybean cell suspension cultures

Abstract

A 3- O-methyltransferase which catalyzes the methylation of caffeic acid to ferulic acid using S-adenosyl- l-methionine as methyl donor has been isolated and purified about 60-fold from cell suspension cultures of soybean ( Glycine max L., var. Mandarin). The enzyme utilized, in addition to caffeic acid ( K m = 133 μM), 5-hydroxyferulic acid ( K m = 55 μM), 3,4,5-trihydroxy-cinnamic acid ( K m = 100 μM), and protocatechualdehyde ( K m = 50 μM) as substrates. Methylation proceeded only in the meta position. The enzyme was unable to catalyze the methylation of ferulic acid, of ortho-, meta-, and para-coumaric acids, and of the flavonoid compounds quercetin and luteolin. The methylation of caffeic acid and 5-hydroxyferulic acid showed a pH optimum at 6.5–7.0. No stimulation of the reaction velocity was observed when Mg 2+ ions were added. EDTA did not inhibit the reaction. The K m for S-adencsyl- l-methionine was 15 μ m. S-Adenosyl- l-homocysteine was a potent competitive inhibitor of S-adenosyl- l-methionine ( K i = 6.9 μM).