Intersubunit Coupling in the Pore of BK Channels
Intersubunit Coupling in the Pore of BK Channels
复制标题
BK 通道孔中的亚基间耦合
DOI：
10.1074/jbc.m109.027789
复制
发表时间：
2009-08-28
影响因子：
4.8
通讯作者：
Ding, Jiuping
中科院分区：
文献类型：
--
作者：
Wu, Ying;Xiong, Yu;Ding, Jiuping
大电导Ca2+激活K+ (BK)通道门控的结构基础仍然难以捉摸。我们发现，在mSlo1 BK通道的S6跨膜段上，用侧链体积较小的亲水性氨基酸取代Leu-312有利于打开状态。通道对钙和电压的敏感性被一些突变所改变，而被另一些突变完全消除。根据变构模型对结果的解释表明，钙不敏感突变体极大地破坏了相对于开放构象的封闭构象的稳定性，也可能破坏Ca2+或电压传感器与栅极之间的变构耦合。一些ph -315突变也倾向于开放状态，这表明Leu-312和ph -315可能在封闭状态下相互作用，形成一个主要的能量屏障，通道必须克服才能打开。同源性模型和分子动力学模拟进一步支持了Leu-312侧链与ph -315芳香环在相邻亚基上的强偶联（L-F偶联）以保持通道闭合。此外，单通道记录表明，钙不敏感突变体的动力学可以近似表征为两态闭开（C-O）模型，在生理条件下表现出接近100%的打开概率，而单通道电导没有改变。这些发现为理解BK通道孔的结构和门控提供了基础。
The structural basis underlying the gating of large conductance Ca2+-activated K+ (BK) channels remains elusive. We found that substitution of Leu-312 in the S6 transmembrane segment of mSlo1 BK channels with hydrophilic amino acids of smaller side-chain volume favored the open state. The sensitivities of channels to calcium and voltage were modified by some mutations and completely abolished by others. Interpretation of the results in terms of an allosteric model suggests that the calcium-insensitive mutants greatly destabilize the closed relative to the open conformation and may also disrupt the allosteric coupling between Ca2+ or voltage sensors and the gate. Some Phe-315 mutations also favor the open state, suggesting that Leu-312 and Phe-315 may interact in the closed state, forming a major energy barrier that the channel has to overcome to open. Homology modeling and molecular dynamic simulations further support that the side chain of Leu-312 can couple strongly with the aromatic ring of Phe-315 in neighboring subunits (L-F coupling) to maintain the channel closed. Additionally, single-channel recordings indicate that the calcium-insensitive mutants, whose kinetics can be approximately characterized by a two-state closed-open (C-O) model, exhibit nearly 100% open probability under physiological conditions without alterations in single-channel conductance. These findings provide a basis for understanding the structure and gating of the BK channel pore.