Interaction of human 15-lipoxygenase-1 with phosphatidylinositol bisphosphates results in increased enzyme activity.
15-lipoxygenase-1 (15-LO-1) can oxygenate both free fatty acids and fatty acids bound to membrane phospholipids. The regulation of the activity of membrane associated 15-LO-1 is poorly understood. Here we demonstrate that calcium ionophore stimulates the translocation of 15-LO-1 to the plasma membrane in human dendritic cells. In a protein-lipid overlay assay, 15-LO-1 was capable of interacting with several phosphoinositides. In the presence of calcium, addition of phosphatidylinositol-4.5-bisphosphate (PI(4.5)P(2)) or PI(3.4)P(2) to the vesicles containing arachidonic acid, led to the formation of approximately three times more 15-HETE than vesicles without phosphoinositides and up to seven times more 15-HETE than vesicles without both calcium and phosphoinositides. The Vmax was unchanged but the apparent Km of 15-LO-1 towards arachidonic acid was significantly lower in the presence of PI(4.5)P(2) or PI(3.4)P(2) in the vesicles in comparison to vesicles with PC only. Taken together, this report demonstrates that human 15-LO-1 binds to PI(4.5)P(2) and PI(3.4)P(2) and that these phospholipids stimulate enzyme activity in the presence of calcium in a vesicle based assay.
Andersson
Erik
E
Division of Hematology, Department of Medicin, Karolinska Institutet and Karolinska University Hospital, Solna, Sweden.
Schain
Frida
F
Svedling
Märta
M
Claesson
Hans-Erik
HE
Forsell
Pontus K A
PK
D001093:Arachidonate 15-Lipoxygenase / Q000235:genetics / Q000378:metabolism*; D000001:Calcimycin / Q000494:pharmacology; D003713:Dendritic Cells / Q000187:drug effects / Q000378:metabolism; D004533:Egtazic Acid / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D007476:Ionophores / Q000494:pharmacology; D007700:Kinetics; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism* / Q000494:pharmacology; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013347:Subcellular Fractions / Q000201:enzymology
D007476:Ionophores; D018129:Phosphatidylinositol Phosphates; D011994:Recombinant Proteins; D000001:Calcimycin; D004533:Egtazic Acid; D001093:Arachidonate 15-Lipoxygenase