Presence of ectonucleotidases in cultured chromaffin cells: hydrolysis of extracellular adenine nucleotides.
The granular ATP released from chromaffin cells during the secretory response can be hydrolyzed by ectonucleotidases that are present in the plasma membrane of these cells. The ecto-ATPase activity showed a Km for ATP of 250 +/- 18 microM and a VMAX value of 167 +/- 25 nmol/10(6) cells x min (1.67 mumol/mg protein x min) for cultured chromaffin cells, while the ecto-ADPase activity showed a Km value for ADP of 375 +/- 40 microM and a VMAX of 125 +/- 20 nmol/10(6) cells x min (1.25 mumol/mg protein x min). The ecto 5'-nucleotidase activity of cultured chromaffin cells was more specific for the purine nucleotides, AMP and IMP, than for the pirimidine nucleotides, CMP and TMP. The Km for AMP was 55 +/- 5 microM and the VMAX value was 4.3 +/- 0.8 nmol/10(6) cells x min (43 nmol/mg protein x min). The nonhydrolyzable analogs of ADP and ATP, alpha, beta-methylene-adenosine 5'-diphosphate and adenylyl-(beta, gamma-methylene)-diphosphonate were good inhibitors of ecto 5'-nucleotidase activity, the KI values being 73.3 +/- 3.5 nM and 193 +/- 29 nM, respectively. The phosphatidylinositol-specific phospholipase C released the ecto-5'-nucleotidase from the chromaffin cells in culture, thus suggesting an anchorage through phosphatidylinositol to plasma membranes. The presence of ectonucleotidases in chromaffin cells may permit the recycling of the extracellular ATP exocytotically released from these neural cells.
Torres
M
M
Departamento de Bioquimica, Facultad de Veterinaria, Universidad Complutense de Madrid, Spain.
Pintor
J
J
Miras-Portugal
M T
MT
D000227:Adenine Nucleotides / Q000378:metabolism*; D000251:Adenosine Triphosphatases / Q000032:analysis*; D000818:Animals; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D002837:Chromaffin Granules / Q000201:enzymology*; D002838:Chromaffin System / Q000201:enzymology*; D002851:Chromatography, High Pressure Liquid; D006868:Hydrolysis; D007700:Kinetics; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D010727:Phosphoric Diester Hydrolases / Q000494:pharmacology; D013379:Substrate Specificity
D000227:Adenine Nucleotides; D010727:Phosphoric Diester Hydrolases; D054801:Phosphoinositide Phospholipase C; D000251:Adenosine Triphosphatases; C027201:ectoATPase; D043265:Phosphatidylinositol Diacylglycerol-Lyase