Naringenin 7-O-methyltransferase involved in the biosynthesis of the flavanone phytoalexin sakuranetin from rice (Oryza sativa L.).

An inducible S-adenosyl-L-methionine:naringenin 7-O-methyltransferase (NOMT) catalyzing the methylation of naringenin to sakuranetin, a major rice phytoalexin was purified approximately 985-fold from ultraviolet (UV)-irradiated rice leaves. The enzyme is not found in healthy tissues and was purified to a nearly homogeneous preparation in one step using adenosine-agarose affinity chromatography, with 1 g rice leaves (UV-irradiated) as starting material. Gel filtration chromatography resulted in an almost pure enzyme, as evidenced by a major band migrating to a position corresponding to a molecular mass of approximately 41 kDa by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The purified NOMT was strongly inhibited by Mn(2+), Ni(2+), Cu(2+), Zn(2+), Hg(2+), and Cd(2+), and to a low degree by Co(2+), Mg(2+), Ba(2+), Ca(2+) and ethylenediamine tetraacetic acid. The amino acid sequence of a NOMT cyanogen bromide (CNBr)-cleavage peptide was highly homologous to that of a caffeic acid 3-O-methyltransferase from maize, and about 70% of the amino acid sequence was obtained after sequencing the peptides generated by CNBr and/or formic acid hydrolysis. NOMT was also shown to be induced in a time-dependent manner, and purified from rice leaves treated with jasmonic acid and copper chloride.