title: 5-lipoxygenase-activating protein stimulates the utilization of arachidonic acid by 5-lipoxygenase.
abstract: 5-Lipoxygenase (5-LO) and its activating protein (FLAP) are both required for cellular leukotriene (LT) synthesis, with 5-LO catalyzing both the synthesis of (5S)-5-hydroperoxy-6,8,11,14-eicosatetraenoic acid (5-HPETE) from arachidonic acid and the subsequent synthesis of LTA4 from 5-HPETE. We have previously expressed both human 5-LO and human FLAP to high levels in Spodoptera frugiperda (Sf9) insect cells, using recombinant baculoviruses. To study the mechanism by which FLAP activates 5-LO, we compared cellular 5-LO activity in Sf9 cells expressing this enzyme to that in Sf9 cells coexpressing FLAP and 5-LO. In this system, FLAP stimulates the utilization of arachidonic acid by 5-LO as a substrate, and increases the efficiency with which 5-LO converts 5-HPETE to LTA4. LT synthesis in cells coexpressing FLAP and 5-LO is inhibited by 3-[1-(p-chlorophenyl)-5-isopropyl-3-tert-butylthio-1H-indol-2-yl]-2,2- dimethyl-propanoic acid (MK-886), an LT biosynthesis inhibitor which specifically binds to FLAP. These studies in Sf9 cells, together with our recent demonstration that FLAP specifically binds arachidonic acid, suggests that FLAP activates 5-LO by acting as an arachidonic acid transfer protein.
authors.affiliation: Department of Molecular Biology, Merck Frosst Centre for Therapeutic Research, Québec, Canada.
authors.forename: M
authors.initials: M
authors.lastname: Abramovitz
authors.forename: E
authors.initials: E
authors.lastname: Wong
authors.forename: M E
authors.initials: ME
authors.lastname: Cox
authors.forename: C D
authors.initials: CD
authors.lastname: Richardson
authors.forename: C
authors.initials: C
authors.lastname: Li
authors.forename: P J
authors.initials: PJ
authors.lastname: Vickers
mesh_terms: D058927:5-Lipoxygenase-Activating Proteins; D000818:Animals; D001094:Arachidonate 5-Lipoxygenase / Q000378:metabolism*; D016718:Arachidonic Acid / Q000378:metabolism*; D002352:Carrier Proteins / Q000494:pharmacology*; D002460:Cell Line; D004789:Enzyme Activation / Q000187:drug effects; D006801:Humans; D015289:Leukotrienes / Q000096:biosynthesis; D008565:Membrane Proteins / Q000494:pharmacology*; D008954:Models, Biological; D009036:Moths; D009504:Neutrophils / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism / Q000494:pharmacology
chemical_list: D058927:5-Lipoxygenase-Activating Proteins; C548592:ALOX5AP protein, human; D002352:Carrier Proteins; D015289:Leukotrienes; D008565:Membrane Proteins; D011994:Recombinant Proteins; D016718:Arachidonic Acid; D001094:Arachidonate 5-Lipoxygenase