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Florian Schaller, Peter Hennig, and Elmar W. Weiler*
The reduction of 12-oxophytodienoic acid (OPDA) to 3-oxo-- 2(2'[Z]-pentenyl)-cyclopentane-1-octanoic acid is catalyzed by 12-- oxophytodienoate-10,11-reductase (OPR). Analysis of the isomer preference of OPR has indicated that the activity is composed of two isoenzymes exhibiting different stereoselectivities. The two isoforms of OPR have been separated, using protein extracts of Rock Harlequin (Corydalis sempervirens) as the starting material. OPRI, the enzyme reported earlier from the same species and corresponding to the cloned OPR from Arabidopsis, utilized 9R,13R-OPDA > > 9S,1 3R-OPDA but not the 13S-configured isomers, whereas the new activity, OPRII, effectively reduced all four OPDA isomers, including the natural 9S,13S-OPDA (cis-[+]-OPDA). OPRII activity is characterized in detail. The enzyme's enzymatic, biochemical, and immunological properties prove that it is a close relative of OPRI. The roles of OPRI and OPRII in octadecanoid biology are discussed.
Abbreviations: AOC, allene oxide cyclase; AOS, allene oxide synthase; JA, jasmonic acid; OPC-8:0, 3-oxo-2(2'[Z]-pentenyl)cyclopentane-1-octanoic acid; OPDA, 12-oxophytodienoic acid; OPR, 12-oxophytodienoate-10,11-reductase; OYE, old yellow enzyme.
The biosynthesis of octadecanoids, cyclic metabolites derived from the Cls fatty acid a-linolenic acid (Vick and Zimmerman, 1984), proceeds in three phases that occur in separate cellular compartments. In phase I, a-linolenic acid is converted to its 13(S)-hydroperoxide, the substrate for the AOS/AOC reaction yielding OPDA. In green tissues these reactions occur in the chloroplast (Bell et al., 1995; Blee and Joyard, 1996; Laudert et al., 1996; for review, see Weiler, 1997). In phase II, OPDA is reduced to OPC-8:0 by a flavoprotein reductase, OPR (EC 1.3.1.42), a soluble, cytosolic protein that was characterized for the first time from corn (Vick and Zimmerman, 1986), was later purified to homogeneity from Rock Harlequin (Corydalis sempervirens; Schaller and Weiler, 1997a), and was finally cloned from Arabidopsis (Schaller and Weiler, 1997b). In phase III, OPC-8:0 is subjected to beta-oxidation to yield JA (Vick and Zimmerman, 1984). Since beta-oxidation in plants occurs only in peroxisomes and glyoxysomes, it is believed that phase III of the octadecanoid biosynthetic pathway is associated with these organelles, although this has not yet been proven.
OPR plays an important role in the biosynthesis of octa-- decanoids in that the enzyme is one of the factors that determines the pool size of OPDA and provides the...