The gastric [H,K]ATPase:H+/ATP stoichiometry

• Published in: The Journal of biological chemistry 1982-06, Vol.257 (11), p.6296-6299
• Main Authors: Rabon, E C, McFall, T L, Sachs, G
• Format: Article
• Language: English
• Subjects: Adenosine Triphosphatases - metabolism; Adenosine Triphosphate; Animals; H(+)-K(+)-Exchanging ATPase; Hydrogen-Ion Concentration; Kinetics; Microsomes - enzymology; Stomach - enzymology
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/S0021-9258(20)65138-9

An H+/ATP ratio of 2 for H+ transport was determined from initial rate measurements at pH 6.1 in a purified gastric microsomal fraction containing the [H,K]ATPase. This ratio was independent of external KCl, though the apparent K0.5 for ATP was increased from 10.78 +/- 0.51 (n = 3) to 64.6 +/- 11.9 (n = 3) microM ATP and from 5.13 +/- 0.64 (n = 3) to 65.2 +/- 0.64 (n = 3) microM ATP for H+ transport and the K+-stimulated ATPase, respectively, as K+external was increased from 12 to 150 mM. The H+/ATP ratio was also relatively independent of ATP concentration. Maximum initial rates obtained in KCl-equilibrated vesicles were independent of added valinomycin, though net H+ transport was increased 29.3 +/- 1.03% (n = 6) by the addition of ionophore. Maximum net H+ transport in this vesicle preparation was 185 +/- 2.1 (n = 14) nmol mg-1 of protein. Initial rate measurements of ATPase represent a burst of K+-dependent activity of approximately 10-15 s duration. The H+/ATP stoichiometry was calculated based on the K+-stimulated component of hydrolysis. Under most conditions, the Mg2+-dependent component of hydrolysis was less than 10% of the (Mg2+ + K+) component of hydrolysis.