Structural Changes in BKCa Channels During Gating

描述（由申请人提供）：大电导电压和Ca2+激活的K+通道（BK）是在控制平滑肌张力和神经元兴奋性中发挥重要作用的膜蛋白。在大多数组织中，它们形成由成孔α亚基和调节亚基组成的复合物。类似地，其他电压依赖性离子通道BK是主要由S4跨膜区段代表的电压传感器。跨膜电位的变化使电压传感器移位，从而产生蛋白质的构象变化。最终，对于足够的去极化，随之而来的构象变化使通道进入允许离子传导的状态。关于调节BK通道开放和关闭的结构知之甚少，并且不存在关于由膜电位变化在BK通道中产生的动态重排的信息。本项目的目的之一是调查BK渠道运作背后的结构变化。a和B亚基的构象变化将通过通道蛋白的残基特异性荧光标记来评估。短期的目标是获得一个更现实的BK蛋白的观点，通过识别区域的运动，电压传感的基础上，这是耦合到激活，失活和失活，以及相对静态的区域，参与其他通道功能。此外，还将研究孔区带电残基的作用。BK通道激活剂目前正在密切研究用于治疗尿失禁和作为中风神经保护剂。这项研究将有助于建立一个框架，设计新的治疗药物或改善已经通过的医疗实践。

DESCRIPTION (provided by applicant): Large conductance Voltage and Ca2+ activated K+ channels (BK) are membrane proteins that play a fundamental role in controlling smooth muscle tone and neuronal excitability. In most of the tissues, they form complexes composed by the pore-forming alpha subunit and by regulatory subunits. Similarly the other voltage dependent ion channels, BK posses a voltage sensor that is mainly represented by the S4 transmembrane segment. Changes in potential across the membrane displace the voltage sensor, producing a conformational change of the protein. Eventually, for adequate depolarizations, the consequent conformational change brings the channel into a state that allows ion conduction. Very little is known about structures that regulate the opening and closing of BK channels, and no information exists about the dynamical rearrangements produced in BK channels by changes in the membrane potential. One of the aims of this project is to investigate the structural changes underlying the operation of BK channel. Conformational changes of both a and b subunits will be assessed by residue specific fluorescent labeling of the channel protein. The short-term objective is to obtain a more realistic view of BK protein by identifying regions of motion that underlie voltage sensing, and that are couple to activation, inactivation and deactivation, and regions of relative staticity, involved in other channel functions. In addition the role of charged residues in pore region will be investigated. BK channel activators are now under close investigation for treatment of urinary incontinence and as stroke neuroprotectant. This study will contribute to set a framework for the design of new therapeutic agents or for the amelioration of the one already adopted by the medical practice.