Regioselectivity of a plant lauric acid omega hydroxylase. Omega hydroxylation of cis and trans unsaturated lauric acid analogs and epoxygenation of the terminal olefin by plant cytochrome P-450.
The study of the stereochemistry and regioselectivity of plant fatty acid hydroxylases is hampered by the difficulty to purify plant cytochrome P-450 enzymes. To provide an alternative, we have now defined an experimental plant system which expresses only one hydroxylase activity towards lauric acid: microsomes from clofibrate-induced Vicia sativa seedlings hydroxylate this fatty acid exclusively at the methyl terminus. To explore the catalytic capabilities of this laurate oxidase, a series of 1-14C-radiolabeled unsaturated lauric acid analogs (7-, 8-, 9-, 10- and 11-dodecenoic acids) were synthesized. Microsomes from clofibrate induced Vicia sativa seedling catalyzed the omega-oxidation of the lauric acid analogs in the presence of O2 and NADPH. The cis and trans forms of the four in-chain unsaturated analogs of lauric acid were 12-hydroxylated with similar efficiency. The terminal olefin was readily converted to the epoxide with only marginal autocatalytic inactivation of the enzyme. The formation of each metabolite was inhibited to the same extend when microsomes were incubated in presence of carbon monoxide or a suicide-substrate for omega LAH, suggesting that a single cytochrome P-450 isoenzyme from Vicia sativa microsomes is able to omega hydroxylate lauric acid and in-chain unsaturated analogs, and to epoxygenate 11-dodecenoic acid.
Weissbart
D
D
Laboratoire d'Enzymologie Cellulaire et Moléculaire, Université Louis Pasteur-CNRS URA 1182, Strasbourg, France.
Salaün
J P
JP
Durst
F
F
Pflieger
P
P
Mioskowski
C
C
D000475:Alkenes / Q000378:metabolism; D042926:Cytochrome P-450 CYP4A; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004852:Epoxy Compounds / Q000378:metabolism; D005229:Fatty Acids, Monounsaturated / Q000494:pharmacology; D007850:Lauric Acids / Q000378:metabolism; D006899:Mixed Function Oxygenases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010944:Plants / Q000201:enzymology*; D013237:Stereoisomerism
D000475:Alkenes; D065607:Cytochrome P-450 Enzyme Inhibitors; D004852:Epoxy Compounds; D005229:Fatty Acids, Monounsaturated; D007850:Lauric Acids; C054728:11-dodecenoic acid; D003577:Cytochrome P-450 Enzyme System; D006899:Mixed Function Oxygenases; D042926:Cytochrome P-450 CYP4A