Stored Ca 2 Depletion-induced Oligomerization of Stromal Interaction Molecule 1 ( STIM 1 ) via the EF-SAM Region AN INITIATION MECHANISM FOR CAPACITIVE Ca 2 ENTRY *
Stored Ca 2 Depletion-induced Oligomerization of Stromal Interaction Molecule 1 ( STIM 1 ) via the EF-SAM Region AN INITIATION MECHANISM FOR CAPACITIVE Ca 2 ENTRY *
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发表时间：
2006
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通讯作者：
P. Stathopulos;Guang-yao Li;M. Plevin;J. Ames;M. Ikura
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作者：
P. Stathopulos;Guang-yao Li;M. Plevin;J. Ames;M. Ikura
基质相互作用分子1（Stromal interaction molecule 1，STIM 1）是钙库调控的钙离子进入的关键分子。内质网（ER）luminalCa 2耗竭导致STIM 1从ER膜同质性重新分布到质膜附近的明显局部聚集体;这些变化先于并与通过Ca 2释放激活通道（CRACs）的细胞质Ca 2内流有关。启动ER STIM 1再分布和质膜CRAC活性的分子机制尚未完全了解。我们重组表达了由EF-手和不育基序（SAM）结构域组成的STIM 1的Ca 2感应区（EF-SAM），以研究其Ca 2相关的构象和生化特征。我们证明，钙负载EFSAM（全息）含有高螺旋度，而EF-SAM在没有钙（载脂蛋白）是紧凑得多。因此，全形物的熔融温度（Tm）比辅基形物高25 °C;热和尿素衍生的热力学参数指示3.2 kcal mol-1的Ca 2诱导的稳定化。我们表明，holoEF-SAM作为单体存在，而apoEFSAM容易形成二聚体和/或低聚物，低聚物到单体的转换，反之亦然，至少部分介导的表面疏水性的变化。此外，我们发现EF-SAM的Ca 2结合亲和力相对较低，表观解离常数（Kd）为0.2-0.6 mM，结合化学计量比为1。我们的研究结果表明，EF-SAM积极参与，并可能是启动STIM 1斑点形成通过大的构象变化的分子触发器。EF-SAM的低Ca 2亲和力与STIM 1作为ER Ca 2传感器的确认作用相一致。
Stromal interaction molecule 1 (STIM1) has recently been identified as a key player in store-operated Ca2 entry. Endoplasmic reticulum (ER) luminalCa2 depletion results in STIM1 redistribution from ER membrane homogeneity to distinctly localized aggregates near the plasma membrane; these changes precede and are linked to cytoplasmic Ca2 influx via Ca2 release-activated channels (CRACs). The molecular mechanisms initiating ER STIM1 redistribution and plasma membrane CRAC activity are not well understood. We recombinantly expressed the Ca2 -sensing region of STIM1 consisting of the EF-hand together with the sterile -motif (SAM) domain (EF-SAM) to investigate its Ca2 -related conformational and biochemical features. We demonstrate that Ca2 -loaded EFSAM (holo) contains high -helicity, whereas EF-SAM in the absence of Ca2 (apo) is much less compact. Accordingly, the melting temperature (Tm) of the holoform is 25 °C higher than apoform; heat and urea-derived thermodynamic parameters indicate a Ca2 -induced stabilization of 3.2 kcal mol 1. We show that holoEF-SAM exists as a monomer, whereas apoEFSAM readily forms a dimer and/or oligomer, and that oligomer to monomer transitions and vice versa are at least in part mediated by changes in surface hydrophobicity. Additionally, we find that the Ca2 binding affinity of EF-SAM is relatively low with an apparent dissociation constant (Kd) of 0.2–0.6 mM and a binding stoichiometry of 1. Our results suggest that EF-SAM actively participates in and is the likely the molecular trigger initiating STIM1 punctae formation via large conformational changes. The lowCa2 affinity of EF-SAM is reconciledwith the confirmed role of STIM1 as an ER Ca2 sensor.