Structure of the CLC-1 chloride channel from Homo sapiens.
Structure of the CLC-1 chloride channel from Homo sapiens.
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DOI：
10.7554/elife.36629
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发表时间：
2018-05-29
期刊：
影响因子：
7.7
通讯作者：
MacKinnon R
中科院分区：
文献类型：
--
作者：
Park E;MacKinnon R
CLC通道介导被动Cl−传导，而CLC转运蛋白介导主动Cl−转运与相反方向的H+转运偶联。CLC-0/1/2通道和CLC转运蛋白之间的区别似乎无法通过氨基酸序列检测到。为了理解为什么它们在功能上不同，我们确定了人类CLC-1通道的结构。它的“谷氨酸门”残基，已知介导CLC转运蛋白中的质子转移，在结构中的位置似乎排除了它的转运功能。此外，较小的侧链在细胞内表面附近产生更宽的孔，可能降低Cl−传导的动力学屏障。当相应的残基在转运蛋白中发生突变时，它被转化为通道。最后，与转运蛋白相比，孔中关键位点的Cl−似乎与降低的亲和力相互作用。因此，谷氨酸门构象，内部孔径和Cl−亲和力的细微差异区分了CLC通道和转运蛋白。通道和转运蛋白是跨细胞膜转运分子和离子（统称为“底物”）的两类蛋白质。通道在膜中形成孔，并且底物被动地扩散通过。另一方面，转运蛋白主动泵送底物穿过膜，在此过程中消耗能量。因此，通道和转运蛋白以不同的方式工作。通道和转运蛋白通常具有不相关的结构，但在结构相似的蛋白质的同一家族中存在两者的例子很少。例如，CLC蛋白质包括氯离子通道和转运蛋白，它们通过利用来自氢离子在另一个方向流动的能量来在一个方向泵送氯离子。目前尚不清楚为什么一些CLC蛋白质作为通道而另一些是转运蛋白，特别是因为这两者似乎无法根据其氨基酸顺序区分-所有蛋白质的构建块。氨基酸序列的保守性意味着它们在结构上非常相似。那么不同的成员如何执行如此能量不同的过程呢？帕克和麦金农现在表明，这个问题的答案提醒人们自然是多么微妙。事实上，虽然人类CLC通道（称为CLC-1）的结构确实与CLC转运蛋白的结构相似，但一个氨基酸采用了独特的形状，这解释了为什么蛋白质不能作为转运蛋白。这种特定的氨基酸，谷氨酸，是中央的氯离子和氢离子交换CLC转运。Park和MacKinnon表明，它在CLC-1通道中的构象阻止了这种交换，同时使孔开放用于氯离子的被动运输。此外，沿着CLC通道中的离子扩散途径的另外两个氨基酸沿着比它们在CLC转运蛋白中的对应物小，因此允许氯离子更快地扩散通过。最后，Park和MacKinnon还指出，通道不需要一个宽孔：相反，如果内部的化学环境允许，离子仍然可以快速流过一个狭窄的孔。CLC蛋白在人类中扮演着许多重要角色，CLC编码基因的突变是许多遗传性疾病的基础。现在知道这种机制研究可能会或可能不会影响治疗还为时过早，但这些发现可能会引起研究离子传导机制和分子功能进化的科学家的兴趣。
CLC channels mediate passive Cl− conduction, while CLC transporters mediate active Cl− transport coupled to H+ transport in the opposite direction. The distinction between CLC-0/1/2 channels and CLC transporters seems undetectable by amino acid sequence. To understand why they are different functionally we determined the structure of the human CLC-1 channel. Its ‘glutamate gate’ residue, known to mediate proton transfer in CLC transporters, adopts a location in the structure that appears to preclude it from its transport function. Furthermore, smaller side chains produce a wider pore near the intracellular surface, potentially reducing a kinetic barrier for Cl− conduction. When the corresponding residues are mutated in a transporter, it is converted to a channel. Finally, Cl− at key sites in the pore appear to interact with reduced affinity compared to transporters. Thus, subtle differences in glutamate gate conformation, internal pore diameter and Cl− affinity distinguish CLC channels and transporters. Channels and transporters are two classes of proteins that transport molecules and ions – collectively referred to as “substrates” – across cell membranes. Channels form a pore in the membrane and the substrates diffuse through passively. Transporters, on the other hand, actively pump substrates across a membrane, consuming energy in the process. Thus, channels and transporters work in distinct ways. Channels and transporters most often have unrelated structures, but there are rare examples of both existing within the same family of structurally similar proteins. CLC proteins, for example, include both chloride ion channels and transporters that pump chloride ions in one direction by harnessing the energy from hydrogen ions flowing in the other direction. It remains unclear why some CLC proteins work as channels while others are transporters, especially since the two seem indistinguishable on the basis of the order of their amino acids – the building blocks of all proteins. The conservation of the amino acid sequences implies they are structurally very similar. How then can different members perform such energetically distinct processes? Park and MacKinnon now show that the answer to this question serves as a reminder of how subtle nature can be. Indeed, while the structure of a human CLC channel (called CLC-1) is indeed similar to those of CLC transporters, one amino acid adopts a unique shape that explains why the protein cannot act as a transporter. This specific amino acid, a glutamate, is central to the exchange of chloride and hydrogen ions in CLC transporters. Park and MacKinnon show that its conformation in the CLC-1 channel stops this exchange, while leaving the pore open for the passive transport of chloride ions. Also, two other amino acids along the ion diffusion pathway in the CLC channel are smaller than their counterparts in CLC transporters, and so allow chloride ions to diffuse through more quickly. Lastly, Park and MacKinnon also note that channels do not require a wide pore: instead ions can still flow rapidly through a narrow pore if the chemical environment inside permits it. CLC proteins perform a number of important roles in humans, and mutations in CLC-encoding genes underlie numerous heritable diseases. It remains too early to know how this mechanistic study may or may not impact treatments, yet the findings will likely interest scientists working on ion conduction mechanisms and the evolution of molecular function.