Monomeric solubilized sarcoplasmic reticulum Ca pump protein: demonstration of Ca binding and dissociation coupled to ATP hydrolysis.
Monomeric solubilized sarcoplasmic reticulum Ca pump protein: demonstration of Ca binding and dissociation coupled to ATP hydrolysis.
复制标题
单体溶解的肌浆网 Ca 泵蛋白：演示 Ca 结合和解离与 ATP 水解的耦合。
DOI：
10.1073/pnas.81.21.6623
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发表时间：
1984
影响因子：
11.1
通讯作者：
Reynolds,JA
中科院分区：
文献类型：
--
作者：
Martin,DW;Tanford,C;Reynolds,JA
肌浆网钙泵蛋白可以在非离子去污剂中以单体形式溶解，并完全保留ATP酶活性。直接证明ATP水解和Ca2+转运之间的耦合保持在可溶状态是不可能的，因为需要单独的隔室Ca2+摄取和Ca2+排放来证明这一点，但在这里，我们提供了强有力的间接证据，耦合实际上持续存在，在正常泵循环的正向和反向方向。正向偶联的证明利用了溶解的蛋白质在没有结合的Ca2+的情况下结构不稳定的事实。伴随ATP水解在溶液中的活性损失是定量一致的顺序Ca2+结合和解离在水解周期。ATP合成对Ca 2+浓度的依赖性证明了反向耦合。虽然溶解（单体）和膜结合蛋白质之间存在实质性差异，如前所述，他们不影响这项工作的主要结论，这是自由能耦合在活性钙离子转运的分子机制是一个固有的特性，每个单独的催化多肽链的泵蛋白。
The sarcoplasmic reticulum Ca-pump protein can be solubilized in monomeric form in nonionic detergents with full retention of ATPase activity. It is impossible to prove directly that coupling between ATP hydrolysis and Ca2+ transport is maintained in the soluble state, because separate compartments for Ca2+ uptake and Ca2+ discharge are required to demonstrate this, but here we provide strong indirect evidence that coupling in fact persists, in both the forward and reverse directions of the normal pump cycle. Demonstration of coupling in the forward direction makes use of the fact that the solubilized protein is structurally labile in the absence of bound Ca2+. Loss of activity accompanying ATP hydrolysis in solution is quantitatively consistent with sequential Ca2+ binding and dissociation during the hydrolysis cycle. Coupling in the reverse direction is demonstrated by the dependence of ATP synthesis on Ca2+ concentration. Although substantial differences between solubilized (monomeric) and membrane-bound protein are shown to exist, as previously reported, they do not affect the main conclusion from this work, which is that the molecular machinery for free-energy coupling in active Ca2+ transport is an inherent property of each individual catalytic polypeptide chain of the pump protein.