Leukotriene formation by a purified reticulocyte lipoxygenase enzyme. Conversion of arachidonic acid and 15-hydroperoxyeicosatetraenoic acid to 14, 15-leukotriene A4.
The purified lipoxygenase of rabbit reticulocytes converts arachidonic acid at 0 degrees C to 15-hydroperoxyeicosatetraenoic acid (15-HPETE) and to 12-hydroperoxyeicosatetraenoic acid (12-HPETE) via reactions which involve hydrogen abstraction at C-13 and C-10, respectively. At 37 degrees C the enzyme converts arachidonic acid to additional products which were identified as 13-hydroxy-14,15-epoxy-5,8,11-eicosatrienoic acid, 8,15-dihydroperoxy-5,9,11,13- and 5,15-dihydroperoxy-6, 6,8,11,13-eicosatetraenoic acids (8,15-diHPETE and 5,15-HPETE, respectively) and diastereoisomers of 8,15-dihydroxy-5,9,11,13-eicosatetraenoic acid (8,15-diHPETEs). The 8,15- and 5,15-diHPETEs were formed by double lipoxygenation since each incorporated 2 molecules of 18O2 and since their synthesis from 15-HPETE was blocked under anaerobic conditions. The 8,15-diHETEs each incorporated 18O from 18O2 at C-15 and were found to arise from nonenzymatic hydrolysis of an epoxytriene which was identified as 14,15-leukotriene A4 by trapping in acidic methanol. This compound was a major product of 15-HPETE in anaerobic incubations. The conversion of 15-HPETE to 14,15-leukotriene A4 was inhibited by the lipoxygenase inhibitors nordihydroguairetic acid and 5,8,11,14-eicosatetraynoic acid. The 14,15-leukotriene A4 synthase and 15-lipoxygenase activities were inhibited by 5,8,11,14-eicosatetraynoic acid in a similar time-dependent manner. The results support a mechanism whereby 14,15-leukotriene A4 is synthesized from 15-HPETE by a further enzymatic step carried out by the reticulocyte 15-lipoxygenase via hydrogen abstraction at C-10 and a redox cycle of the non-heme iron atom of the enzyme.
Bryant
R W
RW
Schewe
T
T
Rapoport
S M
SM
Bailey
J M
JM
D000818:Animals; D001091:Arachidonate Lipoxygenases; D016718:Arachidonic Acid; D001095:Arachidonic Acids / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid; D017572:Leukotriene A4; D015289:Leukotrienes; D008054:Lipid Peroxides / Q000378:metabolism*; D008084:Lipoxygenase / Q000378:metabolism*; D011817:Rabbits; D012156:Reticulocytes / Q000201:enzymology*; D013056:Spectrophotometry, Ultraviolet
D001095:Arachidonic Acids; D017572:Leukotriene A4; D015289:Leukotrienes; D008054:Lipid Peroxides; D016718:Arachidonic Acid; C025086:15-hydroperoxy-5,8,11,13-eicosatetraenoic acid; D001091:Arachidonate Lipoxygenases; D008084:Lipoxygenase