Mechanisms of CLC Transporters and Channels

CLC（“氯离子通道”）家族包含两种主要的离子传输机制：一半 的 CLC 同系物是电扩散离子通道，一半是次级主动转运蛋白 以化学计量将 Cl– 交换为 H 。一个家族中两种机制的发生 表明他们通过共同主题的变体来运作。确实，实验结果支持 CLC 通道是“损坏的”转运蛋白的假设，其中 内、外城门已失。因此，蛋白质构象动力学和离子的细微差别 结合以及它们之间的相互作用可以产生两种不同类型的离子传输 具有相同二级结构的蛋白质的行为。了解其分子基础 这些差异将有助于我们理解 CLC 通道和传输器。 作为二级主动转运蛋白，CLC 转运蛋白利用储存在一个离子中的能量 电化学梯度（Cl– 或 H ）逆着梯度泵送其他离子。这通过发生 蛋白质构象变化与离子结合和运输事件的紧密耦合。发展 CLC 传输机制中离子耦合的完全集成的结构描述，这 该项目将结合互补的尖端方法，包括冷冻电子显微镜 确定高分辨率结构，双电子-电子共振光谱 监测不同条件和不同条件下转运体的构象状态 突变、MD 模拟以确定各种条件下的水合途径，以及 将结构动力学与功能联系起来的定量功能分析。

The CLC (“Chloride Channel”) family encompasses two major ion-transport mechanisms: half of CLC homologs are electrodiffusive ion channels, and half are secondary active transporters that stoichiometrically exchange Cl– for H+. The occurrence of two mechanisms in one family suggests they operate by variations on a common theme. Indeed, experimental results support the hypothesis that CLC channels are “broken” transporters, in which tight coordination between inner and outer gates is lost. Thus, subtle differences in protein conformational dynamics and ion binding—and the interactions between them—can produce two different types of ion transport behavior in proteins with the same secondary structure. Understanding the molecular basis of these differences will inform our understanding of both CLC channels and transporters. As secondary active transporters, CLC transporters harness energy stored in one ion's electrochemical gradient (Cl– or H+) to pump the other ion against its gradient. This occurs through tight coupling of protein conformational changes to ion binding and transport events. To develop a fully integrated structural description of ion coupling in the CLC transport mechanism, this project will combine complementary cutting-edge approaches, including cryo-electron microscopy to determine high-resolution structures, double electron-electron resonance spectroscopy to monitor the conformational state of the transporter under different conditions and with different mutations, MD simulations to determine hydration pathways under various conditions, and quantitative functional assays to connect structural dynamics to function.