ISOLATION OF A MICROSOMAL-ENZYME SYSTEM INVOLVED IN GLUCOSINOLATE BIOSYNTHESIS FROM SEEDLINGS OF TROPAEOLUM-MAJUS L

An in vitro system that converts phenylalanine to phenylacetaldoxime i n the biosynthesis of the glucosinolate glucotropaeolin has been estab lished in seedlings of Tropaeolum majus L. exposed to the combined tre atment of jasmonic acid, ethanol, and light. The treatment resulted in a 9-fold induction, compared with untreated, dark-grown seedlings, of de novo biosynthesis measured as incorporation of radioactively label ed phenylalanine into glucotropaeolin. Formation of the inhibitory deg radation product benzylisothiocyanate during tissue homogenization was prevented by inactivation of the thioglucosidase myrosinase by additi on of 100 mM ascorbic acid to the isolation buffer. This allowed the i solation of a biosynthetically active microsomal preparation from the induced T. majus plant material. The enzyme, which catalyzes the conve rsion of phenylalanine to the corresponding oxime, was sensitive to cy tochrome P450 inhibitors, indicating the involvement of a cytochrome P 450 in the biosynthetic pathway. It has previously been shown that the oxime-producing enzyme in the biosynthesis of p-hydroxybenzylglucosin olate in Sinapis alba L. is dependent on cytochrome P450, whereas the oxime-producing enzymes in Brassica species have been suggested to be flavin monooxygenases or peroxidase-type enzymes. The result with T. m ajus provides additional experimental documentation for a similarity b etween the enzymes converting amino acids into the corresponding oxime s in the biosynthesis of glucosinolates and cyanogenic glucosides.