Reaction mechanism of the gastric H+ +K+-dependent ATPase. Effects of inhibitor and pH.
The effect of nolinium bromide [2-(3,4-dichlorophenylamino)quinolizium bromide], which acts as a K+ antagonist in the gastric H+ +K+-dependent ATPase reaction, was investigated at the level of 32P-labelled intermediates of the gastric ATPase reaction. A concentration-dependent effect of nolinium bromide was observed on the concentrations of phosphorylated intermediates. At low (up to 50 microM) concentrations the drug did not interfere with the concentrations of intermediates but exhibited a competition with K+ at the level of both 32P-labelled intermediates and hydrolysis of ATP at pH 7.0. Similar competition was noted in the H+ +K+-dependent ATPase reaction. Low nolinium bromide concentrations also drastically slowed the enzyme turnover. The concentrations of the intermediates were lowered appreciably between 50 microM- and 100 microM-nolinium bromide without affecting the ATP hydrolysis, and the effects were independent of pH. Similar to the effects at pH 7.0, the drug also exhibited competition with K+ in lowering the E approximately P concentration at pH 5.0. A dramatic effect of pH on the K+-sensitivity as well as on turnover of the 32P-labelled intermediates was observed. Although the concentrations of intermediates remained nearly unaltered at various pH values, the K+-stimulated hydrolysis of ATP showed an optimum at pH 7.0 with sharp declines at pH 5 and 8. The data suggest a critical involvement of H+ in the conversion of the K+-insensitive E1 approximately P into the K+-sensitive E2 approximately P form of the enzyme. Nolinium bromide appears to function as a K+ analogue and seems to block the entry of K+ at the K X E2 step, thereby interfering with the enzyme turnover.
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000814:Aniline Compounds / Q000494:pharmacology; D000818:Animals; D004285:Dogs; D005765:Gastrointestinal Agents / Q000494:pharmacology; D017506:H(+)-K(+)-Exchanging ATPase; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007425:Intracellular Membranes / Q000187:drug effects / Q000201:enzymology; D008861:Microsomes / Q000187:drug effects / Q000201:enzymology; D011807:Quinolizines / Q000494:pharmacology; D013270:Stomach / Q000187:drug effects / Q000201:enzymology*
D000814:Aniline Compounds; D005765:Gastrointestinal Agents; D011807:Quinolizines; D000251:Adenosine Triphosphatases; D017506:H(+)-K(+)-Exchanging ATPase; C019025:nolinium bromide