Divergence of Ca(2+) selectivity and equilibrium Ca(2+) blockade in a Ca(2+) release-activated Ca(2+) channel.
Divergence of Ca(2+) selectivity and equilibrium Ca(2+) blockade in a Ca(2+) release-activated Ca(2+) channel.
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Ca（2+）CA（2+）释放激活Ca（2+）通道中CA（2+）的选择性和平衡Ca（2+）阻滞的发散。
DOI：
10.1085/jgp.201311108
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发表时间：
2014-03
期刊：
影响因子：
--
通讯作者：
Prakriya M
中科院分区：
文献类型：
--
作者：
Yamashita M;Prakriya M
CRAC通道的Ca2+选择性取决于离子进入和退出的动力学以及稳态Ca2+结合亲和力。流行的模型假设，高Ca2+选择性的Ca2+释放激活的Ca2+（CRAC）通道产生紧密的Ca2+结合到孔内的高亲和力位点，从而阻止单价离子通量。在这里，我们研究了高亲和力Ca2+结合对重组Orai3通道中Ca2+选择性的贡献，当由内质网Ca2+传感器STIM 1门控时，Orai3通道作为高Ca2+选择性通道发挥作用，或者当由小分子2-氨基乙氧基二苯基硼酸酯（2-APB）激活时，Orai3通道作为低Ca2+选择性通道发挥作用。在两种门控模式下，细胞外Ca2+以相似的抑制常数（Ki; μ 25 µ M）阻断Na+电流。因此，由Ki的Ca2+阻断所设定的平衡结合不能解释两种门控模式的不同Ca2+选择性。与STIM 1门控通道不同，2-APB门控通道中的Ca 2+阻滞取决于细胞外Na+浓度，并表现出异常陡峭的电压依赖性，与Na+孔占有率的增强一致。此外，在2-APB门控通道中，Ca 2+阻滞的二级速率常数比STIM 1门控通道快8倍。一个四势垒，三个结合位点的Eyring模型表明，降低进入和退出的能量障碍的Ca 2+和Na+模拟2-APB门控通道的速度常数定性再现其低Ca 2+的选择性，表明离子的进入和退出速率强烈影响Ca 2+的选择性。噪声分析表明，2-APB门控通道的单位Na+电导是STIM 1门控通道的四倍，但两种门控模式都显示出较高的开放概率（Po; P <0.7）。在这两种情况下，通道激活过程中电流噪声的增加与闭合通道逐步募集到高Po状态一致，表明潜在的门控机制在两种门控模式下在操作上相似。这些结果表明，高亲和力的Ca 2+结合和动力学因素有助于CRAC通道中的高Ca 2+选择性。
The Ca2+ selectivity of CRAC channels depends on the kinetics of ion entry and exit as well as the steady-state Ca2+ binding affinity. Prevailing models postulate that high Ca2+ selectivity of Ca2+ release-activated Ca2+ (CRAC) channels arises from tight Ca2+ binding to a high affinity site within the pore, thereby blocking monovalent ion flux. Here, we examined the contribution of high affinity Ca2+ binding for Ca2+ selectivity in recombinant Orai3 channels, which function as highly Ca2+-selective channels when gated by the endoplasmic reticulum Ca2+ sensor STIM1 or as poorly Ca2+-selective channels when activated by the small molecule 2-aminoethoxydiphenyl borate (2-APB). Extracellular Ca2+ blocked Na+ currents in both gating modes with a similar inhibition constant (Ki; ∼25 µM). Thus, equilibrium binding as set by the Ki of Ca2+ blockade cannot explain the differing Ca2+ selectivity of the two gating modes. Unlike STIM1-gated channels, Ca2+ blockade in 2-APB–gated channels depended on the extracellular Na+ concentration and exhibited an anomalously steep voltage dependence, consistent with enhanced Na+ pore occupancy. Moreover, the second-order rate constants of Ca2+ blockade were eightfold faster in 2-APB–gated channels than in STIM1-gated channels. A four-barrier, three–binding site Eyring model indicated that lowering the entry and exit energy barriers for Ca2+ and Na+ to simulate the faster rate constants of 2-APB–gated channels qualitatively reproduces their low Ca2+ selectivity, suggesting that ion entry and exit rates strongly affect Ca2+ selectivity. Noise analysis indicated that the unitary Na+ conductance of 2-APB–gated channels is fourfold larger than that of STIM1-gated channels, but both modes of gating show a high open probability (Po; ∼0.7). The increase in current noise during channel activation was consistent with stepwise recruitment of closed channels to a high Po state in both cases, suggesting that the underlying gating mechanisms are operationally similar in the two gating modes. These results suggest that both high affinity Ca2+ binding and kinetic factors contribute to high Ca2+ selectivity in CRAC channels.
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DOI：
10.1126/science.1228757
发表时间：
2012-12-07
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Hou X;Pedi L;Diver MM;Long SB
通讯作者：
Long SB
影响因子：
8.7
作者：
通讯作者：
--
DOI：
10.1073/pnas.1101664108
发表时间：
2011-08-09
影响因子：
11.1
作者：
Hoover, Paul J.;Lewis, Richard S.
通讯作者：
Lewis, Richard S.
DOI：
10.1085/jgp.58.4.413
发表时间：
1971-10
期刊：
The Journal of general physiology
影响因子：
--
作者：
Armstrong CM
通讯作者：
Armstrong CM
DOI：
10.1073/pnas.86.13.5207
发表时间：
1989-07-01
影响因子：
11.1
作者：
FRIEL, DD;TSIEN, RW
通讯作者：
TSIEN, RW