title: Cation-stimulated ATPase activity in purified plasma membranes from tobacco hornworm midgut.
abstract: Purified goblet cell apical membranes from Manduca sexta larval midgut exhibit a specific ATPase activity approx. 20-fold higher than that in the 100 000 X g pellet of a midgut homogenate. The already substantial ATPase activity in this plasma membrane segment is doubled in the presence of 20-50 mM KCl. At ATP concentrations ranging from 0.1 to 3.0 mM, the presence of 20 mM KCl leads to a 10-fold increase in the enzyme's affinity for ATP. ATPase activity is greatest at a pH of approx. 8. In addition to ATP, GTP serves as a substrate, but CTP, ADP, AMP and p-nitrophenyl phosphate do not. Either Mg2+ or Mn2+ is required for activity and cannot be replaced by Ca2+ or Zn2+. The ATPase activity of goblet cell apical membranes is inhibited by neither the typical (Na+ + K+)-ATPase inhibitors, ouabain and orthovanadate, nor by the typical mitochondrial F1F0-ATPase inhibitors, azide and oligomycin. Although 1.5 microM DCCD is ineffective, 150 microM DCCD leads to total inhibition of ATPase activity. The ATPase activity of goblet cell apical membranes is stimulated not only by K+, but also, in order of decreasing effectiveness, by Rb+, Li+, Na+ and even Mg2+. Replacement of Cl- by Br-, F- and HCO3- has less influence than variation of the cations. However, replacement of Cl- by NO3- inhibits strongly this ATPase activity. The ATPase activity described above is characteristic of the alkali metal ion pump containing apical membranes of goblet cells and is not enhanced to a similar degree in other purified midgut epithelial cell plasma membrane segments. Its localization, its broad cation specificity and its insensitivity to ouabain all mimic properties of active ion transport by the lepidopteran midgut and suggest this ATPase as a possible key component of the lepidopteran electrogenic alkali metal ion pump.
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism*; D000818:Animals; D001386:Azides / Q000494:pharmacology; D002462:Cell Membrane / Q000201:enzymology*; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology; D004064:Digestive System / Q000201:enzymology; D007915:Lepidoptera / Q000201:enzymology*; D008274:Magnesium / Q000378:metabolism; D009840:Oligomycins / Q000494:pharmacology; D010042:Ouabain / Q000494:pharmacology; D011189:Potassium Chloride / Q000378:metabolism; D019810:Sodium Azide; D013379:Substrate Specificity
chemical_list: D001386:Azides; D009840:Oligomycins; D004024:Dicyclohexylcarbodiimide; D010042:Ouabain; D011189:Potassium Chloride; D019810:Sodium Azide; D000251:Adenosine Triphosphatases; D008274:Magnesium