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首页> 外文期刊>Journal of Molecular Biology >Construction of a cyclic nucleotide-gated KcsA K+ channel
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Construction of a cyclic nucleotide-gated KcsA K+ channel

机译：环状核苷酸门控的KcsA K +通道的构建

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摘要

The ability of an ion channel to open in response to a defined stimulus is central to its function. In ligand-gated channels, pore opening is conferred through transduction of a conformational change in a gating domain to the helices of the pore. Here, we present the construction of a designed cyclic nucleotide-gated (CNG) channel, named KcsA-CNG, by addition of a prokaryotic cyclic nucleotide-binding domain to a KcsA-derived K+ channel. This channel is functional in lipid bilayers at physiological pH and has the combined properties of both of its parent-derived components. It conducts K+ and is blocked by the K+ channel inhibitors Na+ and agitoxin-2. Channel open times are increased by about two orders of magnitude compared to wild-type KcsA. The average number of open channels increases by similar to 50% upon addition of cAMP. Although the absolute open probabilities are somewhat variable from one channel to the next, the property of cyclic nucleotide sensitivity is very reproducible. An apparent K-d value of similar to 90 nM was estimated. The successful construction of a cyclic nucleotide-gated KcsA K+ channel suggests that it should be possible to produce channels that will respond to novel ligands. (c) 2005 Elsevier Ltd. All rights reserved.
机译：离子通道响应定义的刺激而打开的能力对其功能至关重要。在配体门控的通道中，通过将门控结构域中的构象变化传导至孔的螺旋结构来赋予开孔作用。在这里，我们介绍了一个设计的环状核苷酸门控（CNG）通道，称为KcsA-CNG的构建，方法是将原核环状核苷酸结合域添加到KcsA衍生的K +通道中。该通道在生理pH下在脂质双层中起作用，并具有其亲本衍生组分的组合特性。它传导K +，并被K +通道抑制剂Na +和agitoxin-2阻断。与野生型KcsA相比，渠道开放时间增加了大约两个数量级。添加cAMP后，开放通道的平均数量增加了大约50％。尽管从一个通道到另一个通道的绝对开放概率有所不同，但环状核苷酸敏感性的特性还是非常可重现的。估计的表观K-d值接近90 nM。环状核苷酸门控的KcsA K +通道的成功构建表明，应该有可能产生对新型配体产生响应的通道。 （c）2005 Elsevier Ltd.保留所有权利。

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