tailieunhanh - Báo cáo khoa học: Purification and properties of a new S-adenosyl-Lmethionine:flavonoid 4¢-O-methyltransferase from carnation (Dianthus caryophyllus L.)

A new enzyme,S-adenosyl-L-methionine:flavonoid 4¢-O-methyltransferase (EC )(F 4¢-OMT), has been puri-fied 1 399-fold from the tissues of carnation (Dianthus caryophyllus L). The enzyme, with a molecular mass of 43–45 kDa and a pI of , specifically methylates the hydroxysubstituent in4¢-positionof theflavones, flavanones and isoflavones in the presence ofS-adenosyl-L-methionine. A high affinity for the flavone kaempferol was observed (Km¼;Vmax¼ )1 Æmg )1 ), while other 4¢-hydroxylated flavonoids proved likewise to be suitable substrates | Eur. J. Biochem. 270 3422-3431 2003 FEBS 2003 doi Purification and properties of a new 5-adenosyl-L-methionineflavonoid 4 -ỡ-methyltransferase from carnation Dianthus caryophyllus L. Paolo Curir1 Virginia Lanzotti2. Marcello Dolci3 Paola Dolci3 Carlo Pasini1 and Gordon Tollin4 1Istituto Sperimentale per la Floricoltura Corso Inglesi 508 Sanremo Italy 2DISTAAM University of Molize Campobasso Italy . University of Torino Grugliasco TO Italy 4Department Biochemistry and Molecular Biophysics University of Arizona Tucson AZ USA A new enzyme S-adenosyl-L-methionine flavonoid 4 -O-methyltransferase EC F 4 -OMT has been purified 1 399-fold from the tissues of carnation Dianthus caryophyllus L . The enzyme with a molecular mass of 43-45 kDa and a pI of specifically methylates the hydroxy substituent in 4 -position of the flavones flavanones and isoflavones in the presence of S-adenosyl-L-methionine. A high affinity for the flavone kaempferol was observed Km M Vmax Limol-miffi -mg 1 . while other 4 -hydroxylated flavonoids proved likewise to be suitable substrates. Enzyme activity had no apparent Mg requirement but was inhibited by SH-group reagents. The optimum pH value for F 4 -OMT activity was found to be around neutrality. Kinetic analysis of the enzyme bi-substrate reaction indicates a Ping-Pong mechanism and excludes the formation of a ternary complex. The F 4 -OMT activity was increased in both in vitro and in vivo carnation tissues by the inoculation with Fusarium oxysporum f. sp. dianthi. The enzyme did not display activity towards hydroxycinnamic acid derivatives some of which are involved as methylated monolignols in lignin biosynthesis the role of this enzyme could be therefore mainly defensive rather than structural although its precise function still needs to be ascertained. Keywords S-adenosyl-L-methionine flavonoid 4 -O-methyl-transferase O-methyltransferase Fusarium oxysporum f. sp. dianthi .