Characterization of Human Liver Leukotriene B4 ω-Hydroxylase P450 (CYP4F2)

We previously reported the cloning of a human liver leukotriene B4 (LTB4) ω-hydroxy-lase P450 designated CYP 4F2 [Kikuta et al. (1994) FEBS Lett. 348, 70–74]. However, the properties of CYP 4F2 remain poorly defined. The preparation solubilized using n-octyl-β-D-glucopyranoside from microsomes of CYP 4F2-expressing yeast cells catalyzes ω- hydroxylation of LTB4, 6-trans-LTB4, lipoxin A4, 8-hydroxyeicosatetraenoate, 12-hydroxye-icosatetraenoate, and 12-hydroxystearate in the presence of rabbit liver NADPH-P450 reductase. In addition, the enzyme shows ethoxycoumarin O-deethylase and p-nitroanisole O-demethylase activities. The enzyme was purified to apparent electrophoretic homogeneity from yeast cells by sequential chromatography of solubilized microsomes through amino-n-hexyl-Sepharose 4B, DEAE-HPLC, and hydroxylapatite HPLC columns. The final preparation showed a specific content of 11.1 nmol of P450/mg of protein, with an apparent molecular mass of 56.3 kDa. CYP 4F2 was distinguished from the closely homologous CYP 4F3 (human neutrophil LTB4 to-hydroxylase) by its much higher K4 for LTB4, inability to ω-hydroxylate lipoxin B4; and extreme instability.