Kinetics of conformational changes associated with potassium binding to and release from Na+/K(+)-ATPase.
Kinetics of conformational changes associated with potassium binding to and release from Na+/K(+)-ATPase.
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与钾与 Na /K( )-ATP 酶结合和释放相关的构象变化动力学。
DOI：
10.1016/s0005-2736(96)00162-9
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发表时间：
1996
期刊：
影响因子：
--
通讯作者：
Robinson,JD
中科院分区：
文献类型：
--
作者：
Pratap,PR;Palit,A;Grassi-Nemeth,E;Robinson,JD
Na+K+-ATP酶在细胞中的功能是将ATP水解产生的能量耦合到Na+的运输和K+的运输。荧光探针IAF（iodoacetamidofluorescin）与该酶共价结合，报告构象变化而不抑制酶活性。本文介绍了实验用狗肾酶标记的IAF检查动力学的构象变化所造成的添加Na+和K+，测定稳态和停流荧光变化。这些荧光变化的动力学作为两种初始条件下的温度交界点进行检查：（a）高荧光形式的酶（Ehigh）与不同的[K+]快速混合;（B）低荧光形式的酶（Elow）与不同的[ATP]快速混合。这些实验表明：（1）Ehigh → Elow的荧光变化速率常数远大于Elow → Ehigh的荧光变化速率常数，（2）两个跃迁的表观活化自由能（Eaapp）不同（3）在稳态条件下，在无Na+存在的情况下，当ATP加入到Elow（K+）中时，IAF荧光没有变化;（4）Ehigh-Elow转变的表观活化自由能不依赖于[K+]（16.4kcal/mol），但Elow-Ehigh转变的表观活化自由能随[ATP]的增加而增加：（5）Ehigh-Elow转变的表观活化自由能与无ATP时的表观活化自由能（34 kcal/mol）基本相同。这些结果可以解释为：（i）从Elow到Ehigh的过渡。IAF报道了一种构象变化，发生后，K+释放到细胞内侧，这是参与Na+结合;（ii）Eaapp增加与[ATP]，同时增加过渡态的熵。因此，ATP似乎在从Elowto Ehigh过渡期间使酶不稳定。
The Na+K+- ATPase functions in cells to couple energy from the hydrolysis of ATP to the transport Na+out and K+in. The fluorescent probe IAF (iodoacetamidofluorescin) covalently binds to this enzyme, reporting conformational changes without inhibiting enzyme activity. This paper describes experiments using dog kidney enzyme labeled with IAF to examine kinetics of conformational changes resulting from added Na+and K+, measured in terms of steady-state and stopped-flow fluorescence changes. Kinetics of these fluorescence changes were examined as a junction of temperature from two initial conditions: (a) enzyme in the high-fluorescence form (Ehigh) was rapidly mixed with varying [K+]; and (b) enzyme in the low-fluorescence form (Elow) was rapidly mixed with varying [ATP]. These experiments showed: (1) The rate constant for the fluorescence change from Ehighto Elowwas much larger than that for the opposite transition, Elowto Ehigh; (2) the apparent free energy of activation (Eaapp) for the two transitions were different (as estimated from Arrhenius plots); (3) under steady-state conditions, IAF fluorescence did not change when ATP was added to Elow(K+) in the absence of Na+; (4) the apparent free energy of activation was independent of [K+] for the Ehighto Elowtransition (at 16.4 kcal/mol) but increased with [ATP] for the Elowto Ehightransition: (5) Eaappfor the Elowto Ehightransition with 1 mM ATP was approximately the same as that in the absence of ATP (34 kcal/mol). These results can be interpreted as: (i) in the transition from Elowto Ehigh. IAF reported a conformational change that occurred after K+release to the intracellular side and which is involved in Na+binding; (ii) Eaappincreased with [ATP], while increasing the entropy of the transition state. Thus, ATP appeared to destabilize the enzyme during the transition from Elowto Ehigh.
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活性位点荧光素标记的 (Na,K) ATP 酶构象变化的表征
影响因子：
3
作者：
S. Karlish
通讯作者：
S. Karlish
DOI：
10.1016/0005-2736(74)90292-2
发表时间：
1974-01-01
期刊：
BIOCHIMICA ET BIOPHYSICA ACTA
影响因子：
--
作者：
JORGENSEN, PL
通讯作者：
JORGENSEN, PL
Na /K( )-ATPase 的反应顺序：快速动力学测量可区分替代方案。
DOI：
--
发表时间：
1991
期刊：
Biochimica et Biophysica Acta
影响因子：
--
作者：
P. Pratap;J. Robinson;M. Steinberg
通讯作者：
M. Steinberg
Na、K 和 Mg2 共存位点的 (Na K) 依赖性 ATP 酶证据的动力学研究。
DOI：
--
发表时间：
1983
期刊：
Biochimica et Biophysica Acta
影响因子：
--
作者：
J. Robinson
通讯作者：
J. Robinson
通过简单的手动测定来表征肾 (Na K)-ATP 酶中的 Rb 闭塞。
DOI：
10.1016/0005-2736(87)90081-2
发表时间：
1987
期刊：
Biochimica et biophysica acta
影响因子：
--
作者：
M. Shani;R. Goldschleger;S. Karlish
通讯作者：
S. Karlish