Enzymatic properties of the 15-lipoxygenase of human cultured keratinocytes.

The arachidonic acid 15-lipoxygenase or linoleic acid omega-6 lipoxygenase of human neonatal foreskin cultured keratinocytes converts arachidonic acid to 15-hydroxy-eicosatetraenoic acid and linoleic acid to 13-hydroxy-linoleic acid. A mean of 93% of the 15-lipoxygenase activity in sonicates of cultured keratinocytes was recovered in the 400,000 X g supernatant, attesting to the cytosolic localization of this enzyme. Optimal 15-lipoxygenase activity in the 400,000 X g supernatant was expressed at pH 6.7-7.3 and in the presence of calcium at a concentration of 2 mM or higher. Keratinocyte 15-lipoxygenase metabolized arachidonic acid (Km = 10.6 microM) and linoleic acid (Km = 9.5 microM) with similar efficiency. Nordihydroguaiaretic acid and 5,8,11,14-eicosatetraynoic acid both inhibited the conversion of arachidonic acid to 15-HETE with respective 50% inhibitory concentrations of 2.0 microM and 0.9 microM, while ATP, GTP, and cyclic AMP had no effect on activity at pH 6.8-7.2. The enzymatic properties of human keratinocyte 15-lipoxygenase thus resemble those of PMN leukocyte 15-lipoxygenase and the mediators generated may contribute to the regulation of cutaneous sensation and inflammation.