title: Essential requirement of magnesium ion for optimal activity of the NADPH oxidase of guinea pig polymorphonuclear leukocytes.
abstract: The NADPH-dependent O2-(H2O2)-forming oxidase-rich plasma membranes were purified from myristate (MA)-activated polymorphonuclear leukocytes using a Percoll-density gradient method. The specific activity of the enzyme in the plasma membrane fraction was twelve times higher than that in the cells. Studies on the effect of divalent cations and chelators on the O2- and H2O2 generating activity of the oxidase showed that Mg2+, but not Ca2+, enhanced the activity significantly. Zn2+, on the other hand, was slightly inhibitory to the oxidase activity. EDTA markedly inhibited the oxidase activity whereas EGTA enhanced it. The optimal oxidase activity was seen in the presence of mumolar concentrations of Mg2+ and reached a maximum at Mg2+ concentrations of 40-50 microM. The addition of Mg2+ resulted in a decrease in the apparent Km of the oxidase for NADPH from 40 microM to 25 microM and an increase in apparent Vmax by 1.5 times. These results suggest that Mg2+ enhances both NADPH binding and catalytic activities of the oxidase.
mesh_terms: D015720:5'-Nucleotidase; D000818:Animals; D002413:Cations, Divalent; D002462:Cell Membrane / Q000201:enzymology; D004492:Edetic Acid / Q000494:pharmacology; D004533:Egtazic Acid / Q000494:pharmacology; D006168:Guinea Pigs; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology*; D009113:Muramidase / Q000097:blood; D009247:NADH, NADPH Oxidoreductases / Q000097:blood*; D019255:NADPH Oxidases; D009251:NADPH-Ferrihemoprotein Reductase / Q000097:blood; D009504:Neutrophils / Q000201:enzymology*; D009708:Nucleotidases / Q000097:blood; D013481:Superoxides / Q000097:blood
chemical_list: D002413:Cations, Divalent; D013481:Superoxides; D004533:Egtazic Acid; D004492:Edetic Acid; D009247:NADH, NADPH Oxidoreductases; D009251:NADPH-Ferrihemoprotein Reductase; D019255:NADPH Oxidases; D009708:Nucleotidases; D015720:5'-Nucleotidase; D009113:Muramidase; D008274:Magnesium