Membrane enzyme systems molecular size determinations by radiation inactivation

Abstract

Classical radiation target theory is applied to radiation inactivation data on lyophilized membranes to obtain values for the molecular weights of membrane-bound enzymes. A molecular weight of 250 000 is estimated for the ( Na + + K +-activated ATPase (ATP phosphohydrolase, EC 3.6.1.3) of human erythrocyte ghosts, guinea pig kidney cortex microsomes or plasma membrane preparations, and crayfish nerve cord. Other results suggest that the Mg 2+-dependent ATPase and the ( Na + + K +)-ATPase have the same molecular weight. Studies with a K +-stimulated microsomal alkaline phosphatase (orthophosphoric acid monoester phosphohydrolase, EC 3.1.3.1) indicate a molecular weight of 140 000 for this enzyme which is thought to be responsible for the dephosphorylation step in the hydrolysis of ATP via the ATPase system. The volume fraction of the red cell membrane devoted to active cation transport is estimated at 0.002–0.04%. Microsomal and plasma membrane preparations apparently contain the same Na + + K +)-ATPase judge by their similar responses to ouabain inhibition, cation stimulation, and radiation inactivation. Radiation inactivation estimates of the molecular weight of acetylcholinesterase (acetylcholine acetylhydrolase, EC 3.1.1.7) and of mitochondrial ATPase are also presented. Studies with intact red cells and ghosts, irradiated while frozen, suggest that it might be possible to estimate the molecular weight of enzymes in intact cells.