Artigas Pablo, Gadsby David C
Laboratory of Cardiac/Membrane Physiology, The Rockefeller University, New York, New York 10021, USA.
Ann N Y Acad Sci. 2002 Nov;976:31-40. doi: 10.1111/j.1749-6632.2002.tb04711.x.
Ion pumps and exchangers are considered to be different from ion channels for two principal reasons. Ion pumps move ions against, whereas ion channels allow ions to move with, the electrochemical potential gradient, and pumps transport ions relatively slowly, approximately 10(2) s(-1), whereas channels conduct ions rapidly, approximately 10(7) s(-1). However, the latter high rate refers only to the open pore, and yet all ion channels contain at least one gate. Not surprisingly, the conformational changes associated with channel gating occur with kinetics similar to those of ion pumping. Indeed, ion pumps may be viewed as ion channels with two gates, one external to, and the other internal to, the ion binding cavity. The simple operational rule for such a pump is that the two gates should never be open simultaneously; otherwise, the pump would become a channel and conduct dissipative fluxes several orders of magnitude larger than, and in the opposite direction to, the active transport fluxes. Analyses of Na(+) ion movements mediated by the Na(+)/K(+) pump under various conditions have suggested that in at least one, short-lived, conformation of the pump, an ion-channel-like structure, closed at its intracellular end, connects the extracellular solution with the ion binding sites deep in the protein core. Here we use the marine toxin, palytoxin, to act on Na(+)/K(+) pumps in outside-out patches excised from cardiac myocytes and so transform the pumps into nonselective cation channels which we study using macroscopic, and single-channel, recording. We find that gating of the palytoxin-induced channels is regulated by the pump's natural ligands. Thus, external K(+) congeners tend to close, and external Na(+) tends to open, an extracellular gate, whereas ATP acts from the cytoplasmic solution to open an intracellular gate. These gating influences echo the normal ion occlusion and deocclusion reactions that first entrap two extracellular K(+) ions within the interior of the pump (between the two gates) and then release them to the cytoplasmic side in a step accelerated by ATP. These results offer the promise of being able to examine ion occlusion and deocclusion steps at the microscopic level in single Na(+)/K(+) pump molecules.
离子泵和离子交换器被认为与离子通道不同，主要有两个原因。离子泵逆着电化学势梯度移动离子，而离子通道则顺着电化学势梯度允许离子移动；而且泵运输离子相对较慢，约为10² s⁻¹，而通道传导离子很快，约为10⁷ s⁻¹。然而，后者的高速率仅指开放的孔道，而所有离子通道都至少包含一个门控。毫不奇怪，与通道门控相关的构象变化发生的动力学与离子泵的相似。实际上，离子泵可被视为具有两个门控的离子通道，一个在离子结合腔外部，另一个在离子结合腔内部。这种泵的简单操作规则是两个门控绝不应同时打开；否则，泵将变成一个通道，传导耗散通量，其比主动运输通量大几个数量级且方向相反。对在各种条件下由Na⁺/K⁺泵介导的Na⁺离子运动的分析表明，在泵的至少一种短暂存在的构象中，一种类似离子通道的结构，在其细胞内末端关闭，将细胞外溶液与蛋白质核心深处的离子结合位点相连。在这里，我们使用海洋毒素刺尾鱼毒素作用于从心肌细胞切下的外翻膜片上的Na⁺/K⁺泵，从而将泵转化为非选择性阳离子通道，我们使用宏观记录和单通道记录对其进行研究。我们发现刺尾鱼毒素诱导的通道的门控受泵的天然配体调节。因此，外部K⁺同系物倾向于关闭，而外部Na⁺倾向于打开一个细胞外门控，而ATP从细胞质溶液作用以打开一个细胞内门控。这些门控影响呼应了正常的离子封闭和去封闭反应，该反应首先将两个细胞外K⁺离子截留在泵的内部（在两个门控之间），然后在ATP加速的步骤中将它们释放到细胞质一侧。这些结果有望能够在单个Na⁺/K⁺泵分子的微观水平上检查离子封闭和去封闭步骤。
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