title: The possible importance of the cation-binding site for the oxidative modification of liver 5'-nucleotidase.
abstract: The oxidative damage of proteins and lipid peroxidation of membrane lipoproteins has already been described as a possible pathogenic mechanism for liver injury. The aim of the present study was to examine the mechanism that could be responsible for the oxidative modification of rat liver 5'-nucleotidase during exposure to different free radical generating systems: FeCl2/ascorbate, xanthine/xanthine oxidase and H2O2. The level of lipid peroxidation products malondialdehyde (MDA), as well as the level of protein carbonyl groups formation was measured in cells and extracellular medium. The activity of 5'-nucleotidase was linearly decreased in both hepatocytes and extracellular medium after exposure to the FeCl2/ascorbate system indicating that the possible mechanism for oxidative modification could be a metal-binding site of the enzyme. In xanthine/xanthine oxidase system the enzyme activity of hepatocytes had decreased in hepatocytes but increased in the extracellular medium indicating that proteolysis of membrane proteins could he responsible for enzyme release in the extracellular medium. When hepatocytes were exposed to a H2O2 free-radical generating system, the activity of 5'-nucleotidase tended to be decreased in cells and decreased in extracellular medium too, indicating that H2O2 could be less reactive in producing an oxidative modification of the enzyme. In order to support the hypothesis that the cation-binding site can be responsible for oxidative modification of the enzyme, the isolated hepatocytes were preincubated with a Ca(2+)-channel blocker (Verapamil) and then exposed to different radical-generating systems. Verapamil had only a slight effect in potentiating the inhibition in the FeCl2/ascorbate system. This probably means that the cellular cation flux and cation binding may be included as a vulnerable site with the greatest importance in the oxidative modification of 5'-nucleotidase.
authors.affiliation: Institute of Biochemistry, Faculty of Medicine, Nis, Serbia.
authors.forename: G
authors.initials: G
authors.lastname: Kocić
authors.forename: P
authors.initials: P
authors.lastname: Vlahović
authors.forename: D
authors.initials: D
authors.lastname: Pavlović
authors.forename: R
authors.initials: R
authors.lastname: Kocić
authors.forename: T
authors.initials: T
authors.lastname: Cvetković
authors.forename: I
authors.initials: I
authors.lastname: Stojanović
mesh_terms: D015720:5'-Nucleotidase / Q000737:chemistry / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D002121:Calcium Channel Blockers / Q000494:pharmacology; D002412:Cations / Q000378:metabolism*; D002478:Cells, Cultured; D005609:Free Radicals; D015227:Lipid Peroxidation; D008099:Liver / Q000201:enzymology*; D008297:Male; D010084:Oxidation-Reduction; D018384:Oxidative Stress; D051381:Rats; D017208:Rats, Wistar; D014700:Verapamil / Q000494:pharmacology; D019820:Xanthine / Q000378:metabolism; D014969:Xanthine Oxidase / Q000378:metabolism
chemical_list: D002121:Calcium Channel Blockers; D002412:Cations; D005609:Free Radicals; D019820:Xanthine; D014700:Verapamil; D014969:Xanthine Oxidase; D015720:5'-Nucleotidase