A novel rat hepatic clofibrate-inducible cytochrome P450 that is not a lauric acid hydroxylase

Abstract

2-Methoxy-6-[1-methylethyl]naphthalene (MMEN) was hydroxylated in an NADPH-dependent manner to the (ω-1)-alcohol and the ( R)-ω- and ( S)- ω-alcohols by rat hepatic microsomes. ( S)- ω-Hydroxylation was selectively induced 7-fold by clofibrate treatment. Phenobarbital, 3-methyl-cholanthrene, dexamethasone, cholestyramine, and MMEN did not induce this activity to the same extent. Incubation of the racemic ω-alcohols with microsomes isolated from rats resulted in a greater rate of degradation of the ( S)- than the ( R)- ω-alcohol confirming ( S)- ω-hydroxylation to be an initial catalytic event. MMEN and lauric acid were not competitive inhibitors of each other in microsomes from clofibrate-treated rats, indicating the ( S)- ω-MMEN hydroxylase to be a different enzyme from the characterized clofibrate-inducible lauric acid hydroxylases, CYP4A1 and CYP4A3. This was confirmed by the observations that (1) lauric acid hydroxylation was inhibited by 0.02% Tween 20 or Tween 80 and 25 μM capric or myristic acids, whereas ω-MMEN hydroxylation was not, (2) ω-MMEN hydroxylation was inhibited by ketoconazole, cholesterol and acetone, whereas lauric acid hydroxylation was not, and (3) CYP4A1 and CYP4A3 expressed in Hep G2 cells did not catalyze MMEN hydroxylation. Microsomes from the lungs of rabbits treated with progesterone and kidney of untreated rats did not support selective ( S)- ω-MMEN hydroxylation, indicating that this activity is not associated with CYP4A4 or CYP4A2, respectively. Leukotriene B 4(LTB 4) hepatic microsomal hydroxylation was not inhibited by MMEN and microsomes from human neutrophils did not support the reaction. These data identify a hitherto uncharacterized cytochrome P450 which is selectively induced by clofibrate and does not catalyze the ω-hydroxylation of the fatty acids or prostaglandins investigated. It is proposed that the enzyme catalyzing the selective ( S)- ω-hydroxylation of MMEN is a novel rat P450 and that it is either a new member of the CYP4 family or a clofibrate-inducible P450 from another gene family.