Ca2+dependent K+Channels: Allosteric Gating

描述（由申请人提供）：本研究的长期目标 是了解电压、Ca 2+和Mg 2+的分子机制 大电导K+通道（BK通道）的依赖性激活。BK 通道具有道路生理功能，包括调节 神经递质的释放和血管直径的控制。作为 BK通道的这些生理功能的结果是重要的 临床重要性例如，BK通道的异常活动已经被发现。 与动物模型中的高血压相关;它们的活性增加可能 降低缺血再灌注诱发心律失常的发生率。在 BK通道电压的激活引起电压传感器的移动 在通道中，Ca 2+或Mg 2+与通道结合以引起构象变化。 改变通道蛋白以打开激活门。现在， K+通道孔已被解决;蛋白质序列潜在的他激活 门，电压传感器，和Ca 2+结合位点已被确定。 然而，电压传感器运动、Ca 2+或Mg 2+结合的方式是不同的。 与激活栅极的开口耦合的电流仍然未知。直到 阐明了这些耦合的结构和能量基础，电压， Ca 2+和Mg 2+敏感性在各种BK通道中被调节，以帮助 它们的生理功能无法被理解。根据以前的研究， 我们假设，一个结构域的通道蛋白， 在物理上接近激活门（RCK结构域，用于调节 K+通道的电导）在这些耦合中是中心的。最近，X射线 晶体结构的RCK域已被解决。以结构为导向， 数据，我们将使用分子生物学扰乱通道结构， 确定其对Ca 2+、Mg 2+或电压的能量贡献的影响， 使用我们最近开发的电生理学方法打开通道。 我们还将使用蛋白质生物化学和核磁共振的方法 核磁共振光谱（NMR），以映射特定的分子内蛋白质 在通道激活期间可能改变的相互作用， 通道功能这些实验将为理解 各种BK通道如何在生理过程中发挥作用， 用于治疗目的的BK通道上的靶点。他们还将有助于 我们对离子通道门控的理解。

DESCRIPTION (provided by applicant): The long-term objective of this research is to understand the molecular mechanism of the voltage, Ca2+, and Mg2+ dependent activation of large-conductance K+ channels (BK channels). BK channels have road physiological functions, including the modulation of neurotransmitter release and the control of blood vessel diameters. As a consequence of these physiological functions BK channels are of significant clinical importance. For example, abnormal activity of BK channels has been associated with hypertension in animal models; their increased activity may reduce the incidence of ischemia- reperfusion-induced cardiac arrhythmia. In the activation of BK channels voltage induces movements of the voltage sensor in the channel, Ca2+ or Mg2+ binds to the channel to cause conformational changes in the channel protein to open he activation gate. Now the structure of the K+ channel pore has been solved; protein sequences underlying he activation gate, the voltage sensor, and the Ca2+ binding site have been identified. However, the manner n which voltage sensor movements, Ca2+ or Mg2+ binding are coupled to the opening of the activation gate remains unknown. Until the structural and energetic basis of these couplings is elucidated, how voltage, Ca2+ and Mg2+ sensitivities are modulated in various BK channels to subserve their physiological functions cannot be understood. Based on previous studies, we hypothesize that a structural domain of the channel protein that is physically close to the activation gate (the RCK domain for Regulating the conductance of K+ channels) is central in these couplings. Recently, the X-ray crystal structure of the RCK domain has been solved. Guided by the structural data we will perturb the channel structure using molecular biology and determine its impact on the energetic contribution of Ca2+, Mg2+, or voltage to channel opening using our recently developed electrophysiological approaches. We will also use approaches of protein biochemistry and nuclear magnetic resonance spectroscopy (NMR) to map specific intramolecular protein interactions that nay be altered during channel activation and hence control channel function. These experiments will provide a foundation for understanding how various BK channels play their role in physiological processes and define targets on BK channels for therapeutic purposes. They will also contribute to our understanding of ion channel gating in general.