Kinetics of the acid pump in the stomach. Proton transport and hydrolysis of ATP and p-nitrophenyl phosphate by the gastric H,K-ATPase
Hydrolysis of adenosine 5'-triphosphate (ATP) and p-nitrophenyl phosphate by the hydrogen ion-transporting potassium-stimulated adenosine triphosphatase (H,K-ATPase) was investigated. Hydrolysis of ATP was studied at pH 7.4 in vesicles treated with the ionophore nigericin. The kinetic analysis showed negative cooperativity with one high affinity and one low affinity site for ATP. The rate of hydrolysis decreased at 2000 microM ATP indicating a third site for ATP. When the pH was decreased to 6.5 the experimental results followed Michaelis-Menten enzyme kinetics with one low affinity site. Higher concentrations than 750 microM ATP were inhibitory. Proton transport was measured as accumulation of acridine orange in vesicles equilibrated with 150 mM KCl. The transport at various concentrations of ATP in the pH interval from 6.0 to 8.0 correlated well with the Hill equation with a Hill coefficient between 1.5-1.9. The concentration of ATP resulting in half-maximal transport rate increased from 5 microM at pH 6.0 to 420 microM at pH 8.0. At acidic pH the rate of proton transport decreased at 1000 microM ATP. The K+-stimulated p-nitrophenylphosphatase (pNPPase) activity resulted in a Hill coefficient close to 2 indicating cooperative binding of substrate. These kinetic results are used for a further development of the reaction scheme of the H,K-ATPase.
- Research Organization:
- Biomedical Center, Uppsala, Sweden
- OSTI ID:
- 5023560
- Journal Information:
- J. Biol. Chem.; (United States), Vol. 9
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
ATP
HYDROLYSIS
ATP-ASE
BIOCHEMICAL REACTION KINETICS
NITRO COMPOUNDS
PROTONS
MEMBRANE TRANSPORT
MATHEMATICS
MUCOUS MEMBRANES
PH VALUE
PHOSPHATASES
PHOSPHATES
STOMACH
SWINE
ACID ANHYDRASES
ANIMALS
BARYONS
BODY
CHEMICAL REACTIONS
DECOMPOSITION
DIGESTIVE SYSTEM
DOMESTIC ANIMALS
ELEMENTARY PARTICLES
ENZYMES
ESTERASES
FERMIONS
GASTROINTESTINAL TRACT
HADRONS
HYDROLASES
KINETICS
LYSIS
MAMMALS
MEMBRANES
NUCLEONS
NUCLEOTIDES
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORGANS
OXYGEN COMPOUNDS
PHOSPHOHYDROLASES
PHOSPHORUS COMPOUNDS
REACTION KINETICS
SOLVOLYSIS
VERTEBRATES
550201* - Biochemistry- Tracer Techniques