Interactions of 5-lipoxygenase with membranes: studies on the association of soluble enzyme with membranes and alterations in enzyme activity.
Treatment of rat basophilic leukemia cells (RBL-1) with the calcium ionophore A23187 resulted in activation of 5-lipoxygenase, as indicated by an induction of leukotriene release [Orning, L., Hammarström, S., & Samuelsson, B. (1980) Proc. Natl. Acad. Sci. U.S.A. 77, 2017]. The enzyme activation was accompanied by a time-dependent association of 5-lipoxygenase to the particular fraction. When cells were lysed in the presence of 0.05-10 microM CaCl2, the soluble 5-lipoxygenase became associated with the particulate fraction. This was demonstrated by a decrease in immunoreactivities and enzymatic activities in the soluble fraction and a parallel increase in particulate-associated immunoreactivities. The particulate-bound enzyme was not active. Ca2+ induced the membrane association of 5-lipoxygenase when added into the incubation mixtures containing the membrane fraction with either the cytosolic fraction or the purified enzyme. 5-Lipoxygenase also bound to the microsomal-enriched fraction in the presence of Ca2+. Maximal membrane binding was obtained after a 1-min incubation at 4 degrees C. When a fixed amount of isolated membranes (0.2 mg of protein) and increasing cytosolic protein (0.5-4 mg) were used, a linear increase in enzyme binding was observed. The binding became saturated at 3 mg of cytosolic protein/mg of membrane protein. 5-Lipoxygenase binding to the membrane fraction was unaffected by pretreatment of the membranes with trypsin but was inhibited by treating with phospholipase A2, suggesting that phospholipids are involved.(ABSTRACT TRUNCATED AT 250 WORDS)
D000818:Animals; D001094:Arachidonate 5-Lipoxygenase / Q000378:metabolism*; D001091:Arachidonate Lipoxygenases / Q000378:metabolism*; D001491:Basophils / Q000201:enzymology; D000001:Calcimycin / Q000494:pharmacology; D003600:Cytosol / Q000201:enzymology; D004789:Enzyme Activation / Q000187:drug effects; D007700:Kinetics; D007942:Leukemia, Experimental / Q000201:enzymology; D008566:Membranes / Q000201:enzymology; D008861:Microsomes / Q000201:enzymology; D012995:Solubility; D013367:Submitochondrial Particles / Q000201:enzymology; D014407:Tumor Cells, Cultured / Q000201:enzymology
D000001:Calcimycin; D001091:Arachidonate Lipoxygenases; D001094:Arachidonate 5-Lipoxygenase