Regulation of the human leukocyte 5-lipoxygenase: stimulation by micromolar Ca 2+ levels and phosphatidylcholine vesicles

Abstract

Human leukocyte 5-lipoxygenase (EC 1.13.11.12) is unique among the human lipoxygenases not only in its requirement for free ionized calcium, but also in its regulation by a membrane-associated stimulatory factor, the 100000 × g pellet. In the present study, phosphatidylcholine (PC) vesicles, in the absence of 100000 × g pellet, exhibited a dose-dependent stimulatory activity on the 5-lipoxygenase, which was at least as effective as the 100000 × g pellet. Furthermore, the enzyme was activated by isolated human neutrophil plasma membranes and to a lesser degree by endoplasmic reticulum. The chemoattractant peptide fMet-Leu-Phe (0.1 μM), GTP (10 μM), toxin from bacterium Bordetella pertussis (islet activating protein, 5 μg/ml) and their various combinations were unable to modulate the enzymatic activity of the 5-lipoxygenase. Stimulation of the 5-lipoxygenase by relatively low levels of free ionized calcium was observed both in the presence of the pellet and PC vesicles: maximal stimulation was seen at about 10 μM Ca 2+. The human leukocyte leukotriene A 4 synthase activity also exhibited a similar requirement for free calcium ions. The present study indicates that the membrane-associated stimulatory factor of the human leukocyte 5-lipoxygenase may be replaced by PC vesicles. Moreover, the 5-lipoxygenase and leukotriene A 4 synthase activities require significantly lower Ca 2+ levels for maximal activation than has been reported previously.