Half of the (Na+ + K+)-transporting-ATPase-associated K+-stimulated p-nitrophenyl phosphatase activity of gastric epithelial cells is exposed to the surface exterior.
Ouabain inhibited 86RbCl uptake by 80% in rabbit gastric superficial epithelial cells (SEC), revealing the presence of a functional Na+,K+-ATPase [(Na+ + K+)-transporting ATPase] pump. Intact SEC were used to study the ouabain-sensitive Na+,K+-ATPase and K+-pNPPase (K+-stimulated p-nitrophenyl phosphatase) activities before and after lysis. Intact SEC showed no Na+,K+-ATPase and insignificant Mg2+-ATPase activity. However, appreciable K+-pNPPase activity sensitive to ouabain inhibition was demonstrated by localizing its activity to the cell-surface exterior. The lysed SEC, on the other hand, demonstrated both ouabain-sensitive Na+,K+-ATPase and K+-pNPPase activities. Thus the ATP-hydrolytic site of Na+,K+-ATPase faces exclusively the cytosol, whereas the associated K+-pNPPase is distributed equally across the plasma membrane. The study suggests that the cell-exterior-located K+-pNPPase can be used as a convenient and reliable 'in situ' marker for the functional Na+,K+-ATPase system of various isolated cells under noninvasive conditions.
Nandi
J
J
Department of Surgery, State University, New York Health Science Center, Syracuse 13210.
Das
P K
PK
Levine
R A
RA
Ray
T K
TK
D009597:4-Nitrophenylphosphatase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D003911:Dextrans / Q000494:pharmacology; D004847:Epithelial Cells; D004848:Epithelium / Q000201:enzymology; D005665:Furosemide / Q000494:pharmacology; D005753:Gastric Mucosa / Q000187:drug effects / Q000201:enzymology*; D066298:In Vitro Techniques; D009596:Nitrophenols / Q000494:pharmacology; D009943:Organophosphorus Compounds / Q000494:pharmacology; D010042:Ouabain / Q000494:pharmacology; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D011134:Polysaccharides / Q000494:pharmacology; D011188:Potassium / Q000494:pharmacology; D011817:Rabbits; D012414:Rubidium Radioisotopes; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*
D003911:Dextrans; D009596:Nitrophenols; D009943:Organophosphorus Compounds; D011134:Polysaccharides; D012414:Rubidium Radioisotopes; C008644:nitrophenylphosphate; D010042:Ouabain; D005665:Furosemide; D000255:Adenosine Triphosphate; D010744:Phosphoric Monoester Hydrolases; D009597:4-Nitrophenylphosphatase; D000254:Sodium-Potassium-Exchanging ATPase; D011188:Potassium