CHARACTERIZATION OF MG-2+-ATPASE FROM SHEEP KIDNEY MEDULLA - PURIFICATION

Magnesium-dependent adenosine triphosphatase has been purified from sheep kidney medulla plasma membranes. The purification, which is based on treatment of a kidney plasma membrane fraction with 0.5% digitonin in 3 mm MgCl2, effectively separates the Mg2+-ATPase from (Na+ + K+)-ATPase present in the same tissue and yields the Mg2+-ATPase in soluble form. The purified enzyme is activated by a variety of divalent cations and trivalent cations, including Mg2+, Mn2+, Ca2+, Co2+, Fe2+, Zn2+, Eu3+, Gd3+, and VO2+. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the purified enzyme shows two bands with Rf values corresponding to molecular weights of 150,000 and 77,000. The larger peptide is phosphorylated by [γ-32P]ATP, suggesting that this peptide may contain the active site of the Mg2+-ATPase. The Mg2+-ATPase activity is unaffected by the specific (Na+ + K+)-ATPase inhibitor ouabain. © 1979.