title: A pH-dependent superoxide dismutase activity for zinc-free bovine erythrocuprein. Reexamination of the role of zinc in the holoprotein.
abstract: The zinc-free derivative of bovine erythrocuprein, Cu2E2BE, was prepared and its superoxide dismutase activity was measured and compared with that of the holoprotein, Cu2Zn2BE. The dismutase activity of these proteins was measured by quantitating their inhibition of the superoxide-mediated autooxidation of 6-hydroxydopamine, dihydroxyfumaric acid, pyrogallol, and epinephrine. It was found that the superoxide dismutase activity of the zinc-free protein is pH dependent, ranging between 82 +/- 5% (relative to Cu2Zn2BE) at pH 5.8, and 25 +/- 10% at pH 10.2. The overlapping range of assays and buffers verified that these measurements are independent of the method of assay, buffer, and ionic strength (in the range of mu = 0.10 to 0.20). The variation in activity with pH is probably due, at least in part, to the migration of Cu(II) at high pH as described previously [J. S. Valentine, M. W. Pantoliano, P. J. McDonnell, A. R. Burger, and S. J. Lippard, Proc. Natl. Acad. Sci. USA 76, 4245 (1979)], since Cu(II) bound at the zinc binding site has been shown to have little or no dismutase activity. The observation of high activity (82%) for the zinc-free protein at pH 5.8, where Cu(II) is predominantly in the native Cu binding site, and less susceptible to removal by ethylenediaminetetraacetic acid, demonstrates that the presence of Zn(II) in Cu2Zn2BE does not greatly enhance the inherent dismutase activity of Cu(II) in the holoprotein.
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001798:Blood Proteins / Q000378:metabolism*; D002417:Cattle; D004912:Erythrocytes / Q000378:metabolism*; D007700:Kinetics; D008667:Metalloproteins / Q000378:metabolism*; D010084:Oxidation-Reduction; D011487:Protein Conformation; D013482:Superoxide Dismutase / Q000378:metabolism*; D015032:Zinc / Q000494:pharmacology*
chemical_list: D001798:Blood Proteins; D008667:Metalloproteins; D013482:Superoxide Dismutase; D015032:Zinc