Membrane-bound enzymes. III. Protease activity in leucocytes in relation to erythrocyte membranes

Protease activity was detected in membranes of human bovine erythrocytes prepared by the conventional procedures which include washing and removal of the "buffy layer". The enzyme was extracted by 0.75 M KCNS or (NH4)2SO4 and was activated by 0.4 to 0.5 M of the same salts. Colored, particulate hide powder-azure, membrane fractions and soluble proteins such as hemoglobin, casein or albumin were susceptible to hydrolysis by the membraneous protease. Partial purification of the enzyme was accomplished through disc-gel electrophoresis on polyacrylamide in the presence of 0.25% positively charged detergents like cetyltrimethylammonium bromide. An alkaline protease (pH 7.4) with properties similar to those of the erythrocyte enzyme was found in leucocytes. The similarity between the properties of the leucocytic and erythrocytic proteases and the correlation of the activity in erythrocyte membranes with content of white cells in these preparations, suggest that enzymatic activities in the contaminating leucocytes are responsible for the activity of membraneous proteases in erythrocytes.