Pauls H, Bredenbröcker B, Schoner W
Eur J Biochem. 1980 Aug;109(2):523-33. doi: 10.1111/j.1432-1033.1980.tb04824.x.
(Na+ + K+)-ATPase from beef brain and pig kidney are slowly inactivated by chromium(III) complexes of nucleotide triphosphates in the absence of added univalent and divalent cations. The inactivation of (Na+ + K+)-ATPase activity was accompanied by a parallel decrease of the associated K+-activated p-nitrophenylphosphatase and a parallel loss of the capacity to form, Na+-dependently, a phosphointermediate from [gamma-32P]ATP. The kinetics of inactivation and of phosphorylation with [gamma-32P]CrATP and [alpha-32P]CrATP are consistent with the assumption of the formation of a dissociable complex of CrATP with the enzyme (E) followed by phosphorylation of the enzyme: formula: (see text). The dissociation constant of the CrATP complex of the pig kidney enzyme at 37 degrees C was 43 microM. The inactivation rate constant (k + 2 = 0.033 min-1) was in the range of the dissociation rate constant kd of ADP from the enzyme of 0.011 min-1. The phosphoenzyme was unreactive towards ADP as well as to K+. No hydrolysis of the native isolated phosphoenzyme was observed within 6 h under a variety of conditions, but high concentrations of Na+ reactivated it slowly. The capacity of the Cr-phosphoenzyme of 121 +/- 18 pmol/unit enzyme is identical with the capacity of the unmodified enzyme to form, Na+-dependently, a phosphointermediate. The Cr-phosphoenzyme behaved after acid denaturation like an acylphosphate towards hydroxylamine, but the native phosphoenzyme was not affected by it. ATP protected the enzyme against the inactivation by CrATP (dissociation constant of the enzyme ATP complex = 2.5 microM) as well as low concentrations of K+. CrATP was a competitive inhibitor of (Na+ + K+)-ATPase. It is concluded that CrATP is slowly hydrolyzed at the ATP-binding site of (Na+ + K+)-ATPase and inactivates the enzyme by forming an almost non-reactive phosphoprotein at the site otherwise needed for the Na+-dependent proteinkinase reaction as the phosphate acceptor site.
在不添加单价和二价阳离子的情况下，来自牛脑和猪肾的（Na⁺ + K⁺）-ATP酶会被三磷酸核苷酸的铬（III）络合物缓慢灭活。（Na⁺ + K⁺）-ATP酶活性的灭活伴随着相关的K⁺激活的对硝基苯磷酸酶的平行下降，以及从[γ-³²P]ATP以Na⁺依赖方式形成磷酸中间体的能力的平行丧失。用[γ-³²P]CrATP和[α-³²P]CrATP进行灭活和磷酸化的动力学与以下假设一致：CrATP与酶（E）形成可解离的复合物，随后酶发生磷酸化：公式：（见原文）。猪肾酶的CrATP复合物在37℃时的解离常数为43μM。灭活速率常数（k + 2 = 0.033 min⁻¹）在ADP从酶上解离的解离速率常数kd = 0.011 min⁻¹的范围内。磷酸酶对ADP以及K⁺均无反应。在各种条件下，6小时内未观察到天然分离的磷酸酶的水解，但高浓度的Na⁺会使其缓慢重新激活。Cr-磷酸酶的容量为121±18 pmol/单位酶，与未修饰的酶以Na⁺依赖方式形成磷酸中间体的能力相同。酸变性后的Cr-磷酸酶对羟胺表现得像酰基磷酸，但天然磷酸酶不受其影响。ATP可保护酶免受CrATP的灭活（酶-ATP复合物的解离常数 = 2.5μM）以及低浓度K⁺的影响。CrATP是（Na⁺ + K⁺）-ATP酶的竞争性抑制剂。结论是，CrATP在（Na⁺ + K⁺）-ATP酶的ATP结合位点缓慢水解，并通过在Na⁺依赖的蛋白激酶反应所需的位点（作为磷酸受体位点）形成几乎无反应性的磷蛋白来使酶失活。
