PDF(Pubmed)
Abstract：
Voltage-gated calcium channels control key functions of excitable cells, like synaptic transmission in neurons and the contraction of heart and skeletal muscles. To accomplish such diverse functions, different calcium channels activate at different voltages and with distinct kinetics. To identify the molecular mechanisms governing specific voltage sensing properties, we combined structure modeling, mutagenesis, and electrophysiology to analyze the structures, free energy, and transition kinetics of the activated and resting states of two functionally distinct voltage sensing domains (VSDs) of the eukaryotic calcium channel CaV1.1. Both VSDs displayed the typical features of the sliding helix model; however, they greatly differed in ion-pair formation of the outer gating charges. Specifically, stabilization of the activated state enhanced the voltage dependence of activation, while stabilization of resting states slowed the kinetics. This mechanism provides a mechanistic model explaining how specific ion-pair formation in separate VSDs can realize the characteristic gating properties of voltage-gated cation channels.
Communication in our body runs on electricity. Between the exterior and interior of every living cell, there is a difference in electrical charge, or voltage. Rapid changes in this so-called membrane potential activate vital biological processes, ranging from muscle contraction to communication between nerve cells. Ion channels are cellular structures that maintain membrane potential and help ‘excitable’ cells like nerve and muscle cells produce electrical impulses. They are specialized proteins that form highly specific conduction pores in the cell surface. When open, these channels let charged particles (such as calcium ions) through, rapidly altering the electrical potential between the inside and outside the cell. To ensure proper control over this process, most ion channels open in response to specific stimuli, which is known as ‘gating’. For example, voltage-gated calcium channels contain charge-sensing domains that change shape and allow the channel to open once the membrane potential reaches a certain threshold. These channels play important roles in many tissues and, when mutated, can cause severe brain or muscle disease. Although the basic principle of voltage gating is well-known, the properties of individual voltage-gated calcium channels still vary. Different family members open at different voltage levels and at different speeds. Such fine-tuning is thought to be key to their diverse roles in various parts of the body, but the underlying mechanisms are still poorly understood. Here, Fernández-Quintero, El Ghaleb et al. set out to determine how this variation is achieved. The first step was to create a dynamic computer simulation showing the detailed structure of a mammalian voltage-gated calcium channel, called CaV1.1. The simulation was then used to predict the movements of the voltage sensing regions while the channel opened. The computer modelling experiments showed that although the voltage sensors looked superficially similar, they acted differently. The first of the four voltage sensors of the studied calcium channel controlled opening speed. This was driven by shifts in its configuration that caused oppositely charged parts of the protein to sequentially form and break molecular bonds; a process that takes time. In contrast, the fourth sensor, which set the voltage threshold at which the channel opened, did not form these sequential bonds and accordingly reacted fast. Experimental tests in muscle cells that had been engineered to produce channels with mutations in the sensors, confirmed these results. These findings shed new light on the molecular mechanisms that shape the activity of voltage-gated calcium channels. This knowledge will help us understand better how ion channels work, both in healthy tissue and in human disease.
Communication in our body runs on electricity. Between the exterior and interior of every living cell, there is a difference in electrical charge, or voltage. Rapid changes in this so-called membrane potential activate vital biological processes, ranging from muscle contraction to communication between nerve cells. Ion channels are cellular structures that maintain membrane potential and help ‘excitable’ cells like nerve and muscle cells produce electrical impulses. They are specialized proteins that form highly specific conduction pores in the cell surface. When open, these channels let charged particles (such as calcium ions) through, rapidly altering the electrical potential between the inside and outside the cell. To ensure proper control over this process, most ion channels open in response to specific stimuli, which is known as ‘gating’. For example, voltage-gated calcium channels contain charge-sensing domains that change shape and allow the channel to open once the membrane potential reaches a certain threshold. These channels play important roles in many tissues and, when mutated, can cause severe brain or muscle disease. Although the basic principle of voltage gating is well-known, the properties of individual voltage-gated calcium channels still vary. Different family members open at different voltage levels and at different speeds. Such fine-tuning is thought to be key to their diverse roles in various parts of the body, but the underlying mechanisms are still poorly understood. Here, Fernández-Quintero, El Ghaleb et al. set out to determine how this variation is achieved. The first step was to create a dynamic computer simulation showing the detailed structure of a mammalian voltage-gated calcium channel, called CaV1.1. The simulation was then used to predict the movements of the voltage sensing regions while the channel opened. The computer modelling experiments showed that although the voltage sensors looked superficially similar, they acted differently. The first of the four voltage sensors of the studied calcium channel controlled opening speed. This was driven by shifts in its configuration that caused oppositely charged parts of the protein to sequentially form and break molecular bonds; a process that takes time. In contrast, the fourth sensor, which set the voltage threshold at which the channel opened, did not form these sequential bonds and accordingly reacted fast. Experimental tests in muscle cells that had been engineered to produce channels with mutations in the sensors, confirmed these results. These findings shed new light on the molecular mechanisms that shape the activity of voltage-gated calcium channels. This knowledge will help us understand better how ion channels work, both in healthy tissue and in human disease.
摘要：
电压门控钙通道控制兴奋细胞的关键功能，比如神经元的突触传递以及心脏和骨骼肌的收缩。为了完成如此多样化的功能，不同的钙通道在不同的电压和不同的动力学激活。为了确定控制特定电压传感特性的分子机制，我们结合了结构建模，诱变，和电生理学来分析结构，自由能源,真核钙通道CaV1.1的两个功能不同的电压感应域（VSD）的激活和静息状态的转变动力学。两种VSD都显示了滑动螺旋模型的典型特征；但是，它们在外部门控电荷的离子对形成方面存在很大差异。具体来说,激活状态的稳定增强了激活的电压依赖性，而静息状态的稳定减缓了动力学。这种机制提供了一个机械模型，解释了在单独的VSD中形成特定的离子对如何实现电压门控阳离子通道的特征门控特性。
我们体内的交流是靠电进行的。在每个活细胞的外部和内部之间，电荷有差异，或电压。这种所谓的膜电位的快速变化激活了重要的生物过程，从肌肉收缩到神经细胞之间的交流。离子通道是维持膜电位并帮助“兴奋”细胞如神经和肌肉细胞产生电脉冲的细胞结构。它们是在细胞表面形成高度特异性传导孔的特化蛋白质。打开时,这些通道让带电粒子（如钙离子）通过，快速改变细胞内外的电势。为了确保对这一过程的适当控制，大多数离子通道是响应特定刺激而打开的，这被称为“门控”。例如,电压门控钙通道包含电荷感应域，一旦膜电位达到某个阈值，这些电荷感应域就会改变形状并允许通道打开。这些通道在许多组织中发挥重要作用，当变异时，会导致严重的大脑或肌肉疾病。虽然电压门控的基本原理是众所周知的，单个电压门控钙通道的特性仍然不同。不同的家庭成员以不同的电压水平和不同的速度打开。这种微调被认为是他们在身体各个部位扮演不同角色的关键，但是潜在的机制仍然知之甚少。这里,Fernández-Quintero,ElGhaleb等人。着手确定如何实现这种变化。第一步是创建动态计算机模拟，显示哺乳动物电压门控钙通道的详细结构，称为CaV1.1。然后使用模拟来预测当通道打开时电压感测区域的移动。计算机建模实验表明，尽管电压传感器表面上看起来相似，他们的行为不同。最初研讨了钙通道可控的开启速度的四个电压传感器。这是由其构型的变化所驱动的，该结构导致蛋白质的带相反电荷的部分顺序形成并破坏分子键；这个过程需要时间。相比之下,第四个传感器，设置通道打开的电压阈值，没有形成这些顺序债券，因此反应很快。在肌肉细胞中进行的实验测试，这些细胞被设计成产生传感器中突变的通道，证实了这些结果。这些发现为塑造电压门控钙通道活性的分子机制提供了新的思路。这些知识将帮助我们更好地理解离子通道是如何工作的，在健康组织和人类疾病中。
我们体内的交流是靠电进行的。在每个活细胞的外部和内部之间，电荷有差异，或电压。这种所谓的膜电位的快速变化激活了重要的生物过程，从肌肉收缩到神经细胞之间的交流。离子通道是维持膜电位并帮助“兴奋”细胞如神经和肌肉细胞产生电脉冲的细胞结构。它们是在细胞表面形成高度特异性传导孔的特化蛋白质。打开时,这些通道让带电粒子（如钙离子）通过，快速改变细胞内外的电势。为了确保对这一过程的适当控制，大多数离子通道是响应特定刺激而打开的，这被称为“门控”。例如,电压门控钙通道包含电荷感应域，一旦膜电位达到某个阈值，这些电荷感应域就会改变形状并允许通道打开。这些通道在许多组织中发挥重要作用，当变异时，会导致严重的大脑或肌肉疾病。虽然电压门控的基本原理是众所周知的，单个电压门控钙通道的特性仍然不同。不同的家庭成员以不同的电压水平和不同的速度打开。这种微调被认为是他们在身体各个部位扮演不同角色的关键，但是潜在的机制仍然知之甚少。这里,Fernández-Quintero,ElGhaleb等人。着手确定如何实现这种变化。第一步是创建动态计算机模拟，显示哺乳动物电压门控钙通道的详细结构，称为CaV1.1。然后使用模拟来预测当通道打开时电压感测区域的移动。计算机建模实验表明，尽管电压传感器表面上看起来相似，他们的行为不同。最初研讨了钙通道可控的开启速度的四个电压传感器。这是由其构型的变化所驱动的，该结构导致蛋白质的带相反电荷的部分顺序形成并破坏分子键；这个过程需要时间。相比之下,第四个传感器，设置通道打开的电压阈值，没有形成这些顺序债券，因此反应很快。在肌肉细胞中进行的实验测试，这些细胞被设计成产生传感器中突变的通道，证实了这些结果。这些发现为塑造电压门控钙通道活性的分子机制提供了新的思路。这些知识将帮助我们更好地理解离子通道是如何工作的，在健康组织和人类疾病中。
