Characterization and purification of a lipoxygenase inhibitor in human epidermoid carcinoma A431 cells.
A lipoxygenase inhibitor in the cytosolic fraction of human epidermoid carcinoma A431 cells was characterized and purified. The cytosolic inhibitor lost the inhibitory activity upon heating at 75 degrees C for 15 min or pretreating with 1 mg/ml trypsin at 37 degrees C for 60 min. Cytosol, after dialysis, lost the inhibitory activity but its inhibitory activity recovered when 1 mM GSH was added to the dialysate. The inhibitory activity of cytosol was also abolished by treatment either with 1 mM iodoacetate at 4 degrees C for 1 h or with 0.5 mM H2O2. The pI of the inhibitor was approx. 7.0. In addition to 12-lipoxygenase, the inhibitor inhibited the activities of 5-lipoxygenase and fatty acid cyclo-oxygenase in a cell-free system. The inhibitor was purified by a series of column chromatographies using CM Sephadex C-50, Sephadex G-100 SF and Mono P columns. A major 22 kDa protein was obtained that was distinct from selenium-dependent glutathione peroxidase.
Chen
C J
CJ
Department of Pharmacology, College of Medicine, National Cheng Kung University, Tainan, Taiwan 70101, Republic of China.
Huang
H S
HS
Lee
Y T
YT
Yang
C Y
CY
Chang
W C
WC
D016718:Arachidonic Acid / Q000378:metabolism; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003600:Cytosol / Q000737:chemistry / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D005978:Glutathione / Q000494:pharmacology; D006801:Humans; D007461:Iodoacetates / Q000494:pharmacology; D019807:Iodoacetic Acid; D007526:Isoelectric Point; D008084:Lipoxygenase / Q000378:metabolism; D016859:Lipoxygenase Inhibitors / Q000737:chemistry* / Q000302:isolation & purification* / Q000494:pharmacology; D008861:Microsomes / Q000378:metabolism; D008970:Molecular Weight; D013696:Temperature; D014407:Tumor Cells, Cultured
D007461:Iodoacetates; D016859:Lipoxygenase Inhibitors; D016718:Arachidonic Acid; D008084:Lipoxygenase; D005978:Glutathione; D019807:Iodoacetic Acid