Identification of CYP2C9 as a human liver microsomal linoleic acid epoxygenase.
Leukotoxin (9,10-epoxy-12-octadecanoate) and isoleukotoxin (12, 13-epoxy-9-octadecenoate) are monoepoxides of linoleic acid, synthesized by a cytochrome P450 monooxygenase and possibly by an oxidative burst of inflammatory cells. Recent experiments in this laboratory have indicated that the toxicity of leukotoxin and isoleukotoxin is not due to these epoxides, but to the 9,10- and 12, 13-diol metabolites. Leukotoxin and isoleukotoxin are metabolized primarily by the soluble epoxide hydrolase to form leukotoxin diol. Investigations with recombinant cytochrome P450 enzymes have demonstrated that leukotoxin and isoleukotoxin can be formed by these enzymes. This study used a combination of experimental approaches to identify the major cytochrome P450 enzyme in human liver involved in linoleic acid epoxidation. The kinetic paramenters were determined; the K(m) of linoleic acid epoxidation by pooled human liver microsomes was 170 microM and the V(max) was 58 pmol/mg/min. Correlation analysis was performed using individual samples of human liver microsomes, and the best correlation of linoleic acid epoxidation activity was with tolbutamide hydroxylase activity, CYP2C9. Recombinant CYP2C9 was the most active in linoleic acid epoxygenation, and antibody and chemical inhibition also indicated the importance of CYP2C9. This enzyme, therefore, may serve as a therapeutic target in the treatment of inflammation in order to reduce the amount of circulating leukotoxin/isoleukotoxin and their related diols.
Draper
A J
AJ
Department of Chemistry, Bucknell University, Lewisburg, Pennsylvania, 17837, USA.
Hammock
B D
BD
D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D002642:Chick Embryo; D065729:Cytochrome P-450 CYP2C9; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D019787:Linoleic Acid / Q000378:metabolism*; D008041:Linoleic Acids / Q000378:metabolism / Q000633:toxicity; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology*; D011994:Recombinant Proteins / Q000378:metabolism; D039181:Steroid 16-alpha-Hydroxylase; D013250:Steroid Hydroxylases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013379:Substrate Specificity
D065607:Cytochrome P-450 Enzyme Inhibitors; D004791:Enzyme Inhibitors; D008041:Linoleic Acids; D011994:Recombinant Proteins; C051208:9,10-epoxy-12-octadecenoic acid; D003577:Cytochrome P-450 Enzyme System; D019787:Linoleic Acid; D013250:Steroid Hydroxylases; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D001189:Aryl Hydrocarbon Hydroxylases; D039181:Steroid 16-alpha-Hydroxylase