An ecto-nucleotide pyrophosphatase is one of the main enzymes involved in the extracellular metabolism of ATP in rat C6 glioma

The presence of a nucleotide pyrophosphatase (EC 3.6.1.9) on the plasma mem brane of rat C6 glioma has been demonstrated by analysis of the hydrolysis of ATP labeled in the base and in the alpha- and gamma-phosphates. The enzy me degraded ATP into AMP and PPi and, depending on the ATP concentration, a ccounted for similar to 50-75% of the extracellular degradation of ATP. The association of the enzyme with the plasma membrane was confirmed by ATP hy drolysis in the presence of a varying concentration of pyridoxal phosphate- 6-azophenyl-2',4'-disulfonic acid (PPADS), a membrane-impermeable inhibitor of the enzyme. PPADS concentration above 20 mu M abolished the degradation of ATP into AMP and PPi. The nucleotide pyrophosphatase has an alkaline pH optimum and a K-m for ATP of 17 +/- 5 mu M. The enzyme has a broad substra te specificity and hydrolyzes nucleoside triphosphates, nucleoside diphosph ates, dinucleoside polyphosphates, and nucleoside monophosphate esters but is inhibited by nucleoside monophosphates, adenosine 3',5'-bisphosphate, an d PPADS. The substrate specificity characterizes the enzyme as a nucleotide pyrophosphatase/phosphodiesterase I (PD-I). Immunoblotting and autoadenyly lation identified the enzyme as a plasma cell differentiation antigen-relat ed protein. Hydrolysis of ATP terminates the autophosphorylation of a nucle oside diphosphate kinase (NDPK/nm23) detected in the conditioned medium of C6 cultures. A function of the pyrophosphatase/PD-l and NDPK in the puriner gic and pyrimidinergic signal transduction in C6 is discussed.