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Keywords:

  • Divalent metal ions;
  • H+-ATPase;
  • plasma membrane;
  • transport;
  • zinc

summary

Effects of divalent metal ions (Zn, Mg, Mn), metal-ATP complexes, and pH on the activity of the plasma membrane H+-ATPase were investigated. Both native vesicles, isolated by sucrose gradient centrifugation, and solubilized H+-ATPase, purified by linear glycerol gradient centrifugation were used. Zn, as well as Mg and Mn, stimulates the activity of H+-ATPase. Computer calculations of equilibrium concentrations have been used to separate the effects of individual ion complexes and free ionic forms on the enzyme activity. The calculations indicated that Zn as ZnATP+ serves as a substrate of the plasma membrane H+-ATPase. The possibility of serving as substrate depends on the Zn/ATP ratio and the pH of the medium. The native H+-ATPase was activated by divalent metal ions in the order of Mg > Mn > Zn at pH 6.5 and in the order of Zn > Mn > Mg at pH 5.5 for total metal ion concentrations not higher than 3.0 mM. A Zn: ATP ratio of 0.6 at pH 5.5 gave more than 100% of the Mg-dependent ATPase activity (Mg: ATP ratio of 1, pH 5.5). By contrast, free Zn2+ is an effective inhibitor of ATP hydrolysis and H+ transport with a strong dependence upon the pH of the medium. At pH 55 a total ZnSO4 concentration of 1 mM induced no inhibition of ATP hydrolysis, whereas at pH 6.5 the same Zn concentration decreased the activity of the native and purified H+-ATPase to 49% and 34% (control activity: Mg: ATP ratio of 1, no ZnSO4), respectively. We show that both activation by ZnATP2− and inhibition by free Zn2+ of the native plasma membrane H+-ATPase and the partially purified enzyme were strongly affected by the pH of the medium. The results provide evidence that Zn, in combination with the pH of the cytoplasm could play a role in regulating the plasma membrane H+-ATPase and thus in nutrient uptake of plant systems.