MOLECULAR CHARACTERIZATION AND FUNCTIONAL EXPRESSION OF DIHYDROXYPTEROCARPAN 6A-HYDROXYLASE, AN ENZYME SPECIFIC FOR PTEROCARPANOID PHYTOALEXIN BIOSYNTHESIS IN SOYBEAN (GLYCINE-MAX L.)

Four cytochrome P450-dependent enzymes, among them dihydroxypterocarpa n 6a-hydroxylase (D6aH), are specifically involved in the elicitor-ind ucible biosynthesis of glyceollins, the phytoalexins of soybean, Here we report that CYP93A1 cDNA, which we isolated previously from elicito r-induced soybean cells, codes for a protein with D6aH activity, Analy sis of the catalytic properties of recombinant CYP93A1 expressed in ye ast, its NADPH dependency, stereoselectivity and high substrate affini ty confirmed that D6aH is the physiological function of CYP93A1, It th us represents the first isoflavonoid-specific CYP to be characterized at the molecular level, In elicitor-treated soybean cells producing ph ytoalexins, increases in D6aH activity were correlated with elevated t ranscript levels which indicates that expression of the enzyme is regu lated at the level of transcription. Therefore, CYP93A1 cDNA can be us ed as a specific molecular marker for the inducible defense response a gainst pathogen attach, (C) 1998 Federation of European Biochemical So cieties.