Preparation of lipoproteins containing cation-dependent ATPase

1. 1. Methods for the isolation of Na⁺, K⁺ and Mg²⁺ activated ATPases from a variety of tissues are described. The use of mannitol rather than of sucrose appears to be of decisive importance in some instances such as that of liver. 2. 2. Preparations obtained by this method show extremely high degrees of stimulation in the presence of Na⁺ and K⁺, together with Mg²⁺, when compared to activity in the presence of Mg²⁺ alone. Stimulations of between 500 and 700% are commonly obtained. 3. 3. Upon floatation of the preparations in high-density sucrose solutions, activity was recovered in material at the top of the tubes, implying that the ATPase system is a lipoprotein. 4. 4. Preparations from different tissues exhibit differing sensitivities to strophanthin-G, in agreement with sensitiveness of the parent tissues. In every instance, it can be shown that inhibition by strophanthin-G varies inversely as the K⁺ concentration of the system. 5. 5. It can be shown that the brain lipoprotein splits p-nitrophenyl phosphate reaction. 6. 6. Labelling of the ATPase systems of brain and liver shows that ³²P passes from [³²P]ATP into phosphoproteins in the system. Na⁺ accelerates this passage. In the presence of Na⁺ and K⁺ together, the radioactivity of the phosphoprortein is greatly reduced.