PURIFICATION AND CHARACTERIZATION OF ALLENE OXIDE CYCLASE FROM DRY CORN SEEDS

Allene oxide cyclase (AOC; EC 5.3.99.6) catalyzes the cyclization of 1 2,13(S)-epoxy-9(Z),11,15(Z)-octadecatrienoic acid to 12-oxo-10,15(Z)-p hytodienoic acid, the precursor of jasmonic acid (JA). This soluble en zyme was purified 2000-fold from dry corn (Zea mays L.) kernels to app arent homogeneity. The dimeric protein has a molecular mass of 47 kD. Allene oxide cyclase activity was not affected by divalent ions and wa s not feedback-regulated by its product, 12-oxo-10,15(Z)-phytodienoic acid, or by JA. (+/-)-cis-12,13-Epoxy-9(Z)-octadecenoic acid, a substr ate analog, strongly inhibited the enzyme, with 50% inhibition at 20 m u M. Modification of the inhibitor, such as methylation of the carboxy l group or a shift in the position of the epoxy group, abolished the i nhibitory effect, indicating that both structural elements and their p osition are essential for binding to AOC. Nonsteroidal anti-inflammato ry drugs, which are often used to interfere with JA biosynthesis, did not influence AOC activity. The purified enzyme catalyzed the cyclizat ion of 12,13(S)-epoxy-9(Z),11,15(Z)-octadecatrienoic acid derived from linolenic acid, but not that of 12,13(S)-epoxy-9(Z),11-octadecadienoi c acid derived from linoleic acid.