Sheep uterus dual lipoxygenase in the synthesis of 14, 15-leukotrienes

Abstract

Lipoxygenase was purified to homogeneity from sheep uterus cytosol using a combination of ion exchangers, ammonium sulfate fractionation, and gel filtration. The purified enzyme was found to be a homodimeric protein with monomer molecular weight of 66 kDa. When incubated with arachidonic acid, the enzyme showed two lipoxygenase activities producing both 12- and 15-HPETEs at the optimum pH of 5.5. The relative concentration of 12- and 15-HETEs, however, changed with the pH of the reaction, 12-HETE being higher in the alkaline range and 15-HERE being higher in the acidic range. Furthermore the enzyme showed the expected dual lipoxygenase based 14, 15-LTA4 synthase activity as evidenced by the formation of 8, 15-diHETEs, the hydrolysis products of 14, 15-LTA4. Isolation of 14, 15-LTC4 from the homogenates of sheep uterus gave further evidence on the formation of leukotrienes. This is the first report of the formation of 14, 15-series leukotrienes in mammalian reproductive tissue. © 1994 Academic Press, Inc.