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Mouse liver microsomes treated with octylthioglucoside (OTG-microsomes) were examined for copper-stimulated ATPase activity. The activity was about 1 micromol Pi/mg protein/hr under optimal conditions [300 mM KCl, 3 mm MgSO4, 10 mM GSH, 0.5 micron CuSO4, 3 mM ATP and 50 mM acetate buffer at pH5.0]. A reducing agent such as GSH or dithiothreitol was required for the activity, and removal of Cu+ from the reaction mixture by bathocuporinedisulfonate resulted in a complete loss of copper-stimulated ATPase activity. Vanadate inhibited the copper-stimulated ATPase activity. The OTG-microsomes were phosphorylated in a hydroxylamine-sensitive and copper-stimulated way. Iron used instead of copper also stimulated both ATPase and phosphorylation. These results suggest that microsomes from mouse liver contain copper/iron-stimulated P-type ATPase.

To identify the functional unit of Ca(2+)-ATPase in the sarcoplasmic reticulum, we assessed Ca(2+)-transport activities occurring on sarcoplasmic reticulum membranes with different combinations of active and inactive Ca(2+)-ATPase molecules. We prepared heterodimers, consisting of a native Ca(2+)-ATPase molecule and a Ca(2+)-ATPase molecule inactivated by FITC labelling, by fusing vesicles loaded with each type of Ca(2+)-ATPase. The heterodimers exhibited neither Ca(2+) transport nor ATP hydrolysis, suggesting that Ca(2+) transport by the Ca(2+)-ATPase requires an interaction between functional Ca(2+)-ATPase monomers. This finding implies that the functional unit of the Ca(2+)-ATPase is a dimer.

It is known that the addition of NaCl with oligomycin or ATP stimulates ouabain‐sensitive and K+‐dependent p‐nitrophenylphosphatase (pNPPase) activity of Na+/K+‐ATPase. We investigated the mechanism of the stimulation. The combination of oligomycin and NaCl increased the affinity of pNPPase activity for K+. When the ratio of Na+ to Rb+ was 10 in the presence of oligomycin, Rb+‐binding and pNPPase activity reached a maximal level and Na+ was occluded. Phosphorylation of Na+/K+‐ATPase by p‐nitrophenylphosphate (pNPP) was not affected by oligomycin. Because oligomycin stabilizes the Na+‐occluded E1 state of Na+/K+‐ATPase, it seemed that the Na+‐occluded E1 state increased the affinity of the phosphoenzyme formed from pNPP for K+. On the other hand, the combination of ATP and NaCl also increased the affinity of pNPPase for K+ and activated ATPase activity. Both activities were affected by the ligand conditions. Oligomycin noncompetitively affected the activation of pNPPase by NaCl and ATP. Nonhydrolyzable ATP analogues could not substitute for ATP. As NaE1P, which is the high‐energy phosphoenzyme formed from ATP with Na+, is also the Na+‐occluded E1 state, it is suggested that the Na+‐occluded E1 state increases the affinity of the phosphoenzyme from pNPP for K+ through the interaction between α subunits. Therefore, membrane‐bound Na+/K+‐ATPase would function as at least an (αβ)2‐diprotomer with interacting α subunits at the phosphorylation step.