Non-charged amino acids from three different domains contribute to link agonist binding to channel gating in α7 nicotinic acetylcholine receptors
Non-charged amino acids from three different domains contribute to link agonist binding to channel gating in α7 nicotinic acetylcholine receptors
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DOI：
10.1111/j.1471-4159.2007.04771.x
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发表时间：
2007-10-01
影响因子：
4.7
通讯作者：
Criado, Manuel
中科院分区：
文献类型：
--
作者：
Aldea, Marcos;Mulet, Jose;Criado, Manuel
激动剂与烟碱乙酰胆碱受体的结合导致通道开放。以前，我们已经表明，在三个不同的结构域的α 7烟碱受体的几个带电残基参与耦合结合和门控，可能通过静电相互作用的网络。然而，这个网络也可以被其他残基整合。为了检验这一假设，不带电荷的氨基酸突变和突变受体的表达水平和电生理反应进行了测定。位置Asn 47和GIn 48（环2）、11030、Trp 134和GIn 140（环7）以及Thr 264（M2-M3接头）的突变体显示出较差或无效的功能反应，尽管有显著的膜表达。相比之下，突变体F137 A和S265 A表现出功能增益效应。在所有情况下，剂量-反应关系的变化都很小，EC 50值在三倍小到五倍大之间，反对激动剂结合的大修改。除两个受体外，所有受体的峰值电流以相同的速率衰减，不包括对脱敏的大影响。因此，观察到的变化可能主要是由门控特性的改变引起的。此外，双突变体的分析表明，在这些结构域中的一些残基之间的互连，特别是Gln 48与lle 130，这表明通过这些氨基酸的激动剂结合和通道门控之间的潜在耦合。
Binding of agonists to nicotinic acetylcholine receptors results in channel opening. Previously, we have shown that several charged residues at three different domains of the alpha 7 nicotinic receptor are involved in coupling binding and gating, probably through a network of electrostatic interactions. This network, however, could also be integrated by other residues. To test this hypothesis, non-charged amino acids were mutated and expression levels and electrophysiological responses of mutant receptors were determined. Mutants at positions Asn47 and GIn48 (loop 2), 11030, Trp134, and GIn140 (loop 7), and Thr264 (M2-M3 linker) showed poor or null functional responses, despite significant membrane expression. By contrast, mutants F137A and S265A exhibited a gain of function effect. In all cases, changes in dose-response relationships were small, EC50 values being between threefold smaller and fivefold larger, arguing against large modifications of agonist binding. Peak currents decayed at the same rate in all receptors except two, excluding large effects on desensitization. Thus, the observed changes could be mostly caused by alterations of the gating characteristics. Moreover, analysis of double mutants showed an interconnection between some residues in these domains, especially Gln48 with lle130, suggesting a potential coupling between agonist binding and channel gating through these amino acids.