Characterization of a new plasma membrane-associated ecto-5 '-phosphodiesterase/nucleotide-pyrophosphatase from rat hepatocarcinoma AS-30D cells

We have identified in plasma membrane fractions isolated from rat hepatocar cinoma AS-30D ascites cells three glycoproteins of 125 kDa, 115 kDa and 105 kDa (gp125, gp115 and gp105) which become adenylylated using ATP as substr ate, most readily in the presence of EDTA. The gp115 becomes also phosphory lated. The adenylylation of these tumor glycoproteins was much lower than t hat of a group of analogous adenylylatable glycoproteins (gp130, gp120-gp11 0 dimer and gp100) present in normal rat liver plasma membrane. The tumor g lycoproteins were reversibly O-adenylylated at threonine residues, as was t he case for their normal rat liver counterparts. The tumor gp 115, and the gp 120-gp110 dimer from normal rat liver were both isolated using either AT P-affinity chromatography and/or AMP-affinity chromatography. The gp120-gp1 10 dimer from normal rat liver was identified as the plasma cell differenti ation antigen-1 (PC-1 protein), an ecto-5' phosphodiesterase/nucleotide-pyr ophosphatase (5'-PDE/NPPase). The gp115 from tumor cells also exhibited Zn2 +-stimulated 5'-PDE and NPPase activities in alkaline conditions, although it appears to be distinct from the PC-1 protein. We have determined that th e gp115 is an ecto-enzyme that catalyzes the hydrolysis of extracellular AT P, since its adenylylation and phosphorylation were detected in intact cell s using extracellularly added [alpha-P-32]ATP or [gamma-P-32]ATP, respectiv ely, in the absence of any permeabilizing agent.