Molecular determinants within N terminus of Orai3 protein that control channel activation and gating.
Molecular determinants within N terminus of Orai3 protein that control channel activation and gating.
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DOI：
10.1074/jbc.m111.227546
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发表时间：
2011-09-09
期刊：
影响因子：
--
通讯作者：
Romanin C
中科院分区：
文献类型：
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作者：
Bergsmann J;Derler I;Muik M;Frischauf I;Fahrner M;Pollheimer P;Schwarzinger C;Gruber HJ;Groschner K;Romanin C
STIM 1和奥赖代表了钙释放激活的钙通道的关键组分。奥赖通道的激活需要STIM 1的C末端与奥赖的N和C末端偶联。虽然后者似乎是中央在与STIM 1的相互作用，N末端的作用，特别是保守的区域接近第一个跨膜序列是不太好理解。在这里，我们详细研究了这个保守区域在Orai 3逐步删除的功能作用。通过STIM 1或2-氨基乙氧基二苯基硼酸酯（2-APB）和电流门控特性的Orai 3激活的两种模式的分子决定因素进行了映射。增加的N-末端截短揭示了Orai 3的特异性快速失活的逐渐减少，伴随着与钙调蛋白的结合减少。Orai 3的STIM 1依赖性激活维持，只要该保守的N-末端结构域的第二半存在。进一步的截断消除了它，而Orai 3刺激通过2-APB部分保留。总的来说，N-末端保守区在Orai 3电流门控中发挥多方面的作用，对STIM 1和2-APB刺激的激活具有不同的结构要求。
STIM1 and Orai represent the key components of Ca2+ release-activated Ca2+ channels. Activation of Orai channels requires coupling of the C terminus of STIM1 to the N and C termini of Orai. Although the latter appears to be central in the interaction with STIM1, the role of the N terminus and particularly of the conserved region close to the first transmembrane sequence is less well understood. Here, we investigated in detail the functional role of this conserved region in Orai3 by stepwise deletions. Molecular determinants were mapped for the two modes of Orai3 activation via STIM1 or 2-aminoethoxydiphenyl borate (2-APB) and for current gating characteristics. Increasing N-terminal truncations revealed a progressive decrease of the specific fast inactivation of Orai3 concomitant with diminished binding to calmodulin. STIM1-dependent activation of Orai3 was maintained as long as the second half of this conserved N-terminal domain was present. Further truncations abolished it, whereas Orai3 stimulation via 2-APB was partially retained. In aggregate, the N-terminal conserved region plays a multifaceted role in Orai3 current gating with distinct structural requirements for STIM1- and 2-APB-stimulated activation.