Allosteric mechanisms of CA-activated K channel gating

描述（由申请人提供）：离子通道通过响应各种刺激而打开和关闭（门）的能力来动态调节膜渗透性。在识别和表征参与该门控过程的通道蛋白部分方面已经取得了进展，包括检测刺激的“传感器”和调节通过通道孔的离子流的“门”。然而，关于传感器和门的作用的构象事件和相互作用仍然存在许多问题。全面了解传感器/门通信对于理解通道如何正常工作以及它们如何受到内源性因素、药物或疾病的调节可能至关重要。为了确定传感器和门之间通信的性质，我们研究了 BK 型 Ca2 激活 K 通道 (Slo1) 的功能；一个感应膜电压和细胞内 Ca2 以及各种潜在调节因子（包括细胞内 Mg2、血红素和细胞外 Cu2）的通道。 BK 通道参与重要的生理过程，其功能或表达的破坏与高血压、癫痫和癌症等疾病有关。因此，了解 BK 通道调节可能对了解疾病机制和开发更有效的治疗方案具有直接影响。存在变构机制，通过该机制，传感器的激活和 BK 通道中门的打开似乎代表了彼此强烈影响但可以单独发生的不同事件。 Slo1 提供了一个独特的机会来阐明传感器/门通信的功能特性和分子机制。通过研究 Slo1 对极端刺激的响应并利用该通道的特定动力学和稳态特性，我们能够单独研究传感器激活、通道开放以及传感器与门之间的耦合。这些方法和分析工具将应用于突变型 Slo1 通道，以识别传感器/门通信中涉及的残基和相互作用，并了解 Mg2、血红素和 Cu2 如何调节 BK 通道活性。这些结果不仅对于理解 BK 通道的门控和生理调节很重要，而且对于阐明传感器和门如何耦合以及其他离子通道中的门控如何调节也很重要。

DESCRIPTION (provided by applicant): Ion channels dynamically regulate membrane permeability through their ability to open and close (gate) in response to various stimuli. Progress has been made in identifying and characterizing parts of channel proteins that are involved in this gating process including "sensors" that detect stimuli, and "gates" that regulate the flow of ions through the channel pore. However many questions remain concerning the conformational events and interactions that couple the action of sensors and gates. A comprehensive understanding of sensor/gate communication is likely to be of fundamental importance in understanding how channels normally work and how they are modulated by endogenous factors, drugs, or disease. To determine the nature of communication between sensors and gates we have studied the function of the BK-type Ca2+- activated K+ Channel (Slo1); a channel that senses both membrane voltage and intracellular Ca2+ as well as a variety of potential regulatory factors including intracellular Mg2+, Heme, and extracellular Cu2+. BK channels participate in physiologically important processes, and disruptions in their function or expression have been associated with diseases such as hypertension, epilepsy and cancer. Therefore an understanding of BK channel regulation may have direct implications for understanding disease mechanisms and developing more effective therapeutic options. Allosteric mechanisms exist by which the activation of sensors and the opening of the gate in BK channels appear to represent distinct events that strongly influence each other but can occur in isolation. Slo1 offers a unique opportunity to elucidate the functional properties and molecular mechanisms of sensor/gate communication. By studying the response of Slo1 to extreme stimuli and taking advantage of particular kinetic and steady-state properties of this channel we are able to study, in isolation, sensor activation, channel opening, and the coupling between sensor and gate. These methods and analytical tools will be applied to mutant Slo1 channels to identify residues and interactions that are involved in sensor/gate communication, and to understand how Mg2+, Heme and Cu2+ regulate BK channel activity. The results will be important, not only for understanding the gating and physiological regulation of BK channels, but also for elucidating how sensors and gates are coupled and gating is regulated in other ion channels.