Effects of pH on the interaction of ligands with the (H+ + K+)-ATPase purified from pig gastric mucosa
| Autor: | S. Mårdh, Magnus Ljungström, Felix V. Vega |
|---|---|
| Rok vydání: | 1984 |
| Předmět: |
Molar concentration
Swine ATPase Inorganic chemistry Biophysics Biological Transport Active In Vitro Techniques Ligands Biochemistry Dephosphorylation H(+)-K(+)-Exchanging ATPase Reaction rate constant Adenosine Triphosphate ATP hydrolysis Proton transport Carbonic anhydrase Animals Phosphorylation chemistry.chemical_classification Adenosine Triphosphatases 4-Nitrophenylphosphatase Gastric Juice biology Sodium Cell Biology Hydrogen-Ion Concentration Enzyme chemistry Gastric Mucosa biology.protein Potassium Nuclear chemistry |
| Zdroj: | Biochimica et biophysica acta. 769(1) |
| ISSN: | 0006-3002 |
| Popis: | The effects of K + , Na + and ATP on the gastric (H + + K + )-ATPase were investigated at various pH. The enzyme was phosphorylated by ATP with a pseudo-first-order rate constant of 3650 min −1 at pH 7.4. This rate constant increased to a maximal value of about 7900 min −1 when pH was decreased to 6.0. Alkalinization decreased the rate constant. At pH 8.0 it was 1290 min −1 . Additions of 5 mM K + or Na + , did not change the rate constant at acidic pH, while at neutral or alkaline pH a decrease was observed. Dephosphorylation of phosphoenzyme in lyophilized vesicles was dependent on K + , but not on Na + . Alkaline pH increased the rate of dephosphorylation. K + stimulated the ATPase and p -nitrophenylphosphatase activities. At high concentrations K + was inhibitory. Below pH 7.0 Na + had little or no effect on the ATPase and p -nitrophenylphosphatase, while at alkaline pH, Na + inhibited both activities. The effect of extravesicular pH on transport of H + was investigated. At pH 6.5 the apparent K m for ATP was 2.7 μM and increased little when K + was added extravesicularly. At pH 7.5, millimolar concentrations of K + increased the apparent K m for ATP. Extravesicular K + and Na + inhibited the transport of H + . The inhibition was strongest at alkaline pH and only slight at neutral or acidic pH, suggesting a competition between the alkali metal ions and hydrogen ions at a common binding site on the cytoplasmic side of the membrane. Two H + -producing reactions as possible candidates as physiological regulators of (H + + K + )-ATPase were investigated. Firstly, the hydrolysis of ATP per se, and secondly, the hydration of CO 2 and the subsequent formation of H + and HCO 3 − . The amount of hydrogen ions formed in the ATPase reaction was highest at alkaline pH. The H + /ATP ratio was about 1 at pH 8.0. When CO 2 was added to the reaction medium there was no change in the rate of hydrogen ion transport at pH 7.0, but at pH 8.0 the rate increased 4-times upon the addition of 0.4 mM CO 2 . The results indicate a possible co-operation in the production of acid between the H + + K + -ATPase and a carbonic anhydrase associated with the vesicular membrane. |
| Databáze: | OpenAIRE |
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