title: The binding of a K+ competitive ligand, 2-methyl,8-(phenylmethoxy)imidazo(1,2-a)pyridine 3-acetonitrile, to the gastric (H+ + K+)-ATPase.
abstract: 2-Methyl,8-(phenylmethoxy)imidazo(1,2-a)pyridine 3-acetonitrile (SCH 28080) is a freely reversible K+ site inhibitor of the gastric (H+ + K+)-ATPase. In the presence of 2 mMMgSO4, [14C]SCH 28080 bound saturably to gastric vesicle preparations containing the (H+ + K+)-ATPase and was displaced by lumenal K+. A binding stoichiometry of 2.2 +/- 0.1 mol of SCH 28080/mol of catalytic phosphorylation sites was observed. The affinity of SCH 28080 binding was increased approximately 10-fold (to 45 nM) in the presence of 2 mM ATP. High affinity binding also occurred with 2 microM ATP but not with up to 200 microM D-[beta, gamma-CH2]ATP, suggesting that high affinity binding was to a phosphorylated form of the enzyme. In the presence of ATP, the association rate constant was linearly related to the concentration of SCH 28080. However, the association and dissociation rates of SCH 28080 binding were slow, especially at low temperature (at 1.5 degrees C half-maximal binding of 50 nM SCH 28080 was calculated to occur after 232 s). Binding appeared to be predominantly entropy driven with a high activation energy (40 kJ/mol at 37 degrees C). In the absence of ATP, the association rate constant was not linearly related to the concentration of SCH 28080, suggesting that a conformational change in the enzyme was required before binding could occur.
authors.affiliation: Smith Kline & French (Research) Ltd., Welwyn, Herts, United Kingdom.
authors.forename: D J
authors.initials: DJ
authors.lastname: Keeling
authors.forename: A G
authors.initials: AG
authors.lastname: Taylor
authors.forename: C
authors.initials: C
authors.lastname: Schudt
mesh_terms: D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000818:Animals; D000897:Anti-Ulcer Agents / Q000378:metabolism*; D001665:Binding Sites; D001667:Binding, Competitive; D005753:Gastric Mucosa / Q000201:enzymology*; D017506:H(+)-K(+)-Exchanging ATPase; D007093:Imidazoles / Q000378:metabolism* / Q000494:pharmacology; D007700:Kinetics; D008024:Ligands; D008274:Magnesium / Q000494:pharmacology; D008962:Models, Theoretical; D011188:Potassium / Q000378:metabolism*; D013552:Swine; D013816:Thermodynamics
chemical_list: D000897:Anti-Ulcer Agents; D007093:Imidazoles; D008024:Ligands; C035235:Sch 28080; D000251:Adenosine Triphosphatases; D017506:H(+)-K(+)-Exchanging ATPase; D008274:Magnesium; D011188:Potassium