Differences in the Modulations of the Soluble or Plasma‐Membrane‐Bound 5′‐Nucleotidase

Abstract

The treatment of [MOPC 173 murine plasmocytoma cell] plasma membranes by a French pressure cell in sucrose medium devoid of detergents solubilized 20% of the total protein and 95-100% of 5''-nucleotidase activity. The soluble enzyme was 40-90-fold purified by centrifugation in a sucrose gradient with a 10-20% yield with respect to the original lysate. The purified fraction retained the same high specificity for 5''-AMP (Km = 20 .mu.M) as in the plasma membranes and was enriched in sphingomyelin. Whereas 5''-AMP at high concentration inhibited the membrane-bound enzyme, it had no effect on the solubilized form. The soluble enzyme was stimulated by 2 .times. 10-13 - 2 .times. 10-15 g concanavalin A without any inhibition with higher doses of lectin. The plasma membrane bound stimulated and inhibited 5''-nucleotidase was modulated by concanavalin A concentrations higher than 0.1 .mu.g. Inhibition of activity of the soluble enzyme by antiphosphorylcholine antibodies was not observed with membranes. Regulation of 5''-nucleotidase activity in plasma membranes might be associated with a supramolecular organization.