Inorganic pyrophosphatase (
PPiase) activity was measured in cell fractions of rat, mouse, and human erythrocytes; normal rat liver;
Novikoff hepatoma; Morris 3924A
hepatoma; and mouse Ehrlich and
Sarcoma 37 ascites tumors. Despite high intracellular activities, when precautions were taken to maintain cell integrity, only negligible activities were found on the surface of intact erythrocytes, Novikoff
ascites hepatoma, Ehrlich
carcinoma, and
Sarcoma 37 cells.
Suspensions of intact Ehrlich and
Sarcoma 37 cells exhibited low
PPiase activity, but this was only about 1 to 2% of the intracellular activity and was completely accounted for by activity present in the
suspension medium. It is considered to be due to extrusion of the intracellular
enzyme. Systematic fractionation of subcellular components revealed that 92% of the total
PPiase activity of rat liver and 97.5% of that of
Hepatoma 3924A were in the cytosol. The soluble activity consisted of a major form, accompanied by very low activities of two minor forms, all of which migrate toward the
anode on electrophoresis. About 4.5% of the liver activity was present in the mitochondria in two forms, one remaining at the origin and one migrating toward the cathode. The same cytosolic
isozymes were present in
Hepatoma 3924A, and the cathodic form was present in mitochondria but in much reduced amount. No evidence was obtained for specific
isozymes in nuclei or microsomes. Only negligible
PPiase activities were found in cell membranes isolated by
sucrose gradient centrifugation. These results discount the occurrence of
PPiase activity as an ectoenzyme or in the plasma membrane of these cells and point to the cytosol as the major and mitochondria as a minor locus of intracellular
PPiase activity.