Isolation and some properties of dioxygenase and co-oxidase activities of adult human liver cytosolic lipoxygenase.
The evidence presented here constitutes the first report on the occurrence of lipoxygenase (LO) activity in the adult human liver. LO activity was isolated free of hemoglobin from the whole liver cytosol by affinity chromatography using a concanavalin-A sepharose 4B column, and some properties of its dioxygenase and co-oxidase activities were examined. High-pressure liquid chromatography (HPLC) analyses of arachidonic acid metabolites suggested the presence of 5-, 12-, and 15-LO activities in the human liver. Linoleic acid was converted into 13-hydroperoxyoctadecadienoic acid. The dioxygenase activity with a Vmax value of 1.74 mumoles/min/mg protein and a Km value of 0.48 mM was noted in the presence of different concentrations of linoleic acid at pH 10. The activity was markedly stimulated by the presence of calcium, ATP, hydrogen peroxide, and KCl in the assay medium. Under optimum conditions, all the xenobiotics tested were cooxidized by the enzyme preparations in the presence of linoleic acid. Kinetic data obtained for benzidine oxidation yielded a Km value of 0.53 mM and a Vmax value of 90.9 nmoles/min/mg protein. At present, the significance of these findings in in vivo toxicity of benzidine is unknown. The linoleic acid-dependent dioxygenase and co-oxidase activities were thermolabile and inhibited by micromolar concentrations of several classical LO inhibitors, further confirming the involvement of LO in these reactions.
Roy
S K
SK
Florida Toxicology Research Center, Department of Environmental and Occupational Health, College of Public Health, University of South Florida, Tampa 33612-3805, USA.
Kulkarni
A P
AP
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000328:Adult; D016718:Arachidonic Acid / Q000378:metabolism; D001560:Benzidines / Q000737:chemistry; D002118:Calcium / Q000494:pharmacology; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D003208:Concanavalin A / Q000737:chemistry; D003600:Cytosol / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000633:toxicity; D005260:Female; D006454:Hemoglobins / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D019787:Linoleic Acid; D008041:Linoleic Acids / Q000378:metabolism; D008084:Lipoxygenase / Q000302:isolation & purification / Q000378:metabolism*; D008099:Liver / Q000187:drug effects / Q000201:enzymology*; D008297:Male; D008875:Middle Aged; D010084:Oxidation-Reduction; D011189:Potassium Chloride / Q000494:pharmacology; D015262:Xenobiotics / Q000378:metabolism / Q000633:toxicity
D001560:Benzidines; D004791:Enzyme Inhibitors; D006454:Hemoglobins; D008041:Linoleic Acids; D015262:Xenobiotics; D003208:Concanavalin A; D016718:Arachidonic Acid; C029876:benzidine; D011189:Potassium Chloride; D000255:Adenosine Triphosphate; D019787:Linoleic Acid; D006861:Hydrogen Peroxide; D008084:Lipoxygenase; D002118:Calcium