Redistribution and characterization of (H+ + K+)-ATPase membranes from resting and stimulated gastric parietal cells.
When isolated from resting parietal cells, the majority of the (H+ + K+)-ATPase activity was recovered in the microsomal fraction. These microsomal vesicles demonstrated a low K+ permeability, such that the addition of valinomycin resulted in marked stimulation of (H+ + K+)-ATPase activity, and proton accumulation. When isolated from stimulated parietal cells, the (H+ + K+)-ATPase was redistributed to larger, denser vesicles: stimulation-associated (s.a.) vesicles. S.a. vesicles showed an increased K+ permeability, such that maximal (H+ + K+)-ATPase and proton accumulation activities were observed in low K+ concentrations and no enhancement of activities occurred on the addition of valinomycin. The change in subcellular distribution of (H+ + K+)-ATPase correlated with morphological changes observed with stimulation of parietal cells, the microsomes and s.a. vesicles derived from the intracellular tubulovesicles and the apical plasma membrane, respectively. Total (H+ + K+)-ATPase activity recoverable from stimulated gastric mucosa was 64% of that from resting tissue. Therefore, we tested for latent activity in s.a. vesicles. Permeabilization of s.a. vesicles with octyl glucoside increased (H+ + K+)-ATPase activity by greater than 2-fold. Latent (H+ + K+)-ATPase activity was resistant to highly tryptic conditions (which inactivated all activity in gastric microsomes). About 20% of the non-latent (H+ + K+)-ATPase activity was also resistant to trypsin digestion. We interpret these results as indicating that, of the s.a. vesicles, approx. 55% have a right-side-out orientation and are impermeable to ATP, 10% right-side-out and permeable to ATP, and 35% have an inside-out orientation.
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000818:Animals; D002462:Cell Membrane / Q000201:enzymology; D002498:Centrifugation; D003902:Detergents / Q000494:pharmacology; D005260:Female; D005960:Glucosides / Q000494:pharmacology; D017506:H(+)-K(+)-Exchanging ATPase; D066298:In Vitro Techniques; D008297:Male; D009997:Osmotic Pressure; D010295:Parietal Cells, Gastric / Q000187:drug effects / Q000201:enzymology*; D011522:Protons; D011817:Rabbits; D013347:Subcellular Fractions / Q000201:enzymology; D014357:Trypsin / Q000494:pharmacology
D003902:Detergents; D005960:Glucosides; D011522:Protons; C018619:octyl-beta-D-glucoside; D014357:Trypsin; D000251:Adenosine Triphosphatases; D017506:H(+)-K(+)-Exchanging ATPase