Purified pig kidney ATPase was incubated in 30--160 mM Tris-HCl with various monovalent cations. 130 mM LiCl stimulated a ouabain-sensitive ATP hydrolysis (about 5% of the maximal (Na+ + K) activity), whereas 160 mM Tris-HCl did not stimulate hydrolysis. Similar results were obtained with human red blood cell broken membranes. 2. In the absence of Na+ and with 130 mM LiCl, the ATPase activity as a function of KCl concentration showed an initial slight inhibition (50 micrometer KCl) followed by an activation (maximal at 0.2 mM KCl) and a further inhibition, which was total at mM KCl. In the absence of LiCl, the rate of hydrolysis was not affected by any of the KCl concentrations investigated. 3. The lithium-activation curve for ATPase activity in the absence of both Na+ and K+ had sigmoid characteristics. It also showed a marked dependence on the total LiCl + Tris-HCl concentration, being inhibited at high concentrations. This inhibition was more noticeable at low LiCl concentrations. 4. In the absence of Na+, 130 mM Li+ showed promoted phosphorylation of ATPase from 1 to 3 mM ATP in the presence of Mg2+. In enzyme treated with N-ethylmaleimide, the levels of phosphorylation in Li+-containing solutions, amounted to 40% of those in Na+- and up to 7 times of those in K+-containing solutions. 5. The total (Na+ + K+)-ATPase activity was markedly inhibited at high buffer concentrations (Tris-HCl, Imidazole-HCl and tetramethylammonium-HEPES gave similar results) in cases when either the concentration of Na+ or K+ (or both) was below saturation. On the other hand, the maximal (Na+ + K+)-ATPase activity was not affected (or very slightly) by the buffer concentration. 6. Under standard conditions (Tris-HCl + NaCl = 160 mM) the Na+-activation curve of Na+-ATPase had a steep rise between 0 and 2.5 mM, a fall between 2.5 and 20 mM and a further increase between 20 and 130 mM. With 30 mM Tris-HCl, the curve rose more steeply, inhibition was noticeable at 2.5 mM Na+ and was completed at 5 mM Na+. With Tris-HCl + NaCl = 280 mM, the amount of activation decreased and inhibition at intermediate Na+ concentrations was not detected.
摘要
将纯化的猪肾ATP酶与各种单价阳离子一起在30 - 160 mM Tris - HCl中孵育。130 mM LiCl刺激哇巴因敏感的ATP水解（约为最大（Na + + K）活性的5%），而160 mM Tris - HCl不刺激水解。用人红细胞破碎膜也得到了类似结果。2. 在没有Na + 且存在130 mM LiCl的情况下，ATP酶活性作为KCl浓度的函数显示出最初的轻微抑制（50 micrometer KCl），随后是激活（在0.2 mM KCl时最大），然后是进一步抑制，在mM KCl时完全抑制。在没有LiCl的情况下，水解速率不受所研究的任何KCl浓度的影响。3. 在没有Na + 和K + 的情况下，ATP酶活性的锂激活曲线具有S形特征。它还显示出对总LiCl + Tris - HCl浓度有明显依赖性，在高浓度时受到抑制。这种抑制在低LiCl浓度时更明显。4. 在没有Na + 的情况下，130 mM Li + 在存在Mg2 + 时促进了ATP酶从1到3 mM ATP的磷酸化。在用N - 乙基马来酰亚胺处理的酶中，含Li + 溶液中的磷酸化水平相当于含Na + 溶液中的40%，高达含K + 溶液中的7倍。5. 当Na + 或K + （或两者）的浓度低于饱和时，在高缓冲液浓度下（Tris - HCl、咪唑 - HCl和四甲基铵 - HEPES给出类似结果），总（Na + + K + ）-ATP酶活性受到明显抑制。另一方面，最大（Na + + K + ）-ATP酶活性不受（或非常轻微地受）缓冲液浓度影响。6. 在标准条件下（Tris - HCl + NaCl = 160 mM），Na + -ATP酶的Na + 激活曲线在0至2.5 mM之间急剧上升，在2.5至20 mM之间下降，在20至130 mM之间进一步增加。在30 mM Tris - HCl时，曲线上升更陡峭，在2.5 mM Na + 时抑制明显，在5 mM Na + 时完全抑制。在Tris - HCl + NaCl = 280 mM时，激活量减少，在中等Na + 浓度下未检测到抑制。
