Van der Hijden H T, Koster H P, Swarts H G, De Pont J J
Department of Biochemistry, University of Nijmegen, The Netherlands.
Biochim Biophys Acta. 1991 Jan 30;1061(2):141-8. doi: 10.1016/0005-2736(91)90278-g.
The effects of K+ on the phosphorylation of H+/K(+)-ATPase with inorganic phosphate were studied using H+/K(+)-ATPase purified from porcine gastric mucosa. The phosphoenzyme formed by phosphorylation with Pi was identical with the phosphoenzyme formed with ATP. The maximal phosphorylation level obtained with Pi was equal to that obtained with ATP. The Pi phosphorylation reaction of H+/K(+)-ATPase was, like that of Na+/K(+)-ATPase, a relatively slow reaction. The rates of phosphorylation and dephosphorylation were both increased by low concentrations of K+, which resulted in hardly any effect on the phosphorylation level. A decrease of the steady-state phosphorylation level was caused by higher concentrations of K+ in a noncompetitive manner, whereas no further increase in the dephosphorylation rate was observed. The decreasing effect was caused by a slow binding of K+ to the enzyme. All above-mentioned K+ effects were abolished by the specific H+/K(+)-ATPase inhibitor SCH 28080 (2-methyl-8-[phenyl-methoxy]imidazo-[1-2-a]pyrine-3-acetonitrile). Additionally, SCH 28080 caused a 2-fold increase in the affinity of H+/K(+)-ATPase for Pi. A model for the reaction cycle of H+/K(+)-ATPase fitting the data is postulated.
利用从猪胃黏膜中纯化得到的H⁺/K⁺-ATP酶，研究了钾离子（K⁺）对H⁺/K⁺-ATP酶与无机磷酸发生磷酸化作用的影响。由无机磷酸（Pi）磷酸化形成的磷酸化酶与由三磷酸腺苷（ATP）形成的磷酸化酶相同。用Pi获得的最大磷酸化水平与用ATP获得的相同。H⁺/K⁺-ATP酶的Pi磷酸化反应与Na⁺/K⁺-ATP酶的反应一样，是一个相对缓慢的反应。低浓度的K⁺会使磷酸化和去磷酸化的速率都增加，而这对磷酸化水平几乎没有影响。较高浓度的K⁺以非竞争性方式导致稳态磷酸化水平降低，而未观察到去磷酸化速率进一步增加。这种降低作用是由K⁺与酶的缓慢结合引起的。所有上述K⁺的作用都被特异性H⁺/K⁺-ATP酶抑制剂SCH 28080（2-甲基-8-[苯甲氧基]咪唑并-[1,2-a]吡啶-3-乙腈）消除。此外，SCH 28080使H⁺/K⁺-ATP酶对Pi的亲和力增加了两倍。提出了一个符合这些数据的H⁺/K⁺-ATP酶反应循环模型。
