Cymes Gisela D, Grosman Claudio
Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, Urbana, IL 61801.
Department of Molecular and Integrative Physiology, University of Illinois at Urbana-Champaign, Urbana, IL 61801;
Proc Natl Acad Sci U S A. 2016 Nov 8;113(45):E7106-E7115. doi: 10.1073/pnas.1608519113. Epub 2016 Oct 10.
Among neurotransmitter-gated ion channels, the superfamily of pentameric ligand-gated ion channels (pLGICs) is unique in that its members display opposite permeant-ion charge selectivities despite sharing the same structural fold. Although much effort has been devoted to the identification of the mechanism underlying the cation-versus-anion selectivity of these channels, a careful analysis of past work reveals that discrepancies exist, that different explanations for the same phenomenon have often been put forth, and that no consensus view has yet been reached. To elucidate the molecular basis of charge selectivity for the superfamily as a whole, we performed extensive mutagenesis and electrophysiological recordings on six different cation-selective and anion-selective homologs from vertebrate, invertebrate, and bacterial origin. We present compelling evidence for the critical involvement of ionized side chains-whether pore-facing or buried-rather than backbone atoms and propose a mechanism whereby not only their charge sign but also their conformation determines charge selectivity. Insertions, deletions, and residue-to-residue mutations involving nonionizable residues in the intracellular end of the pore seem to affect charge selectivity by changing the rotamer preferences of the ionized side chains in the first turn of the M2 α-helices. We also found that, upon neutralization of the charged residues in the first turn of M2, the control of charge selectivity is handed over to the many other ionized side chains that decorate the pore. This explains the long-standing puzzle as to why the neutralization of the intracellular-mouth glutamates affects charge selectivity to markedly different extents in different cation-selective pLGICs.
在神经递质门控离子通道中，五聚体配体门控离子通道（pLGICs）超家族独具特色，其成员尽管具有相同的结构折叠，但却表现出相反的通透离子电荷选择性。尽管人们付出了诸多努力来确定这些通道阳离子与阴离子选择性背后的机制，但对以往工作的仔细分析表明，存在差异，对于同一现象常常提出不同的解释，且尚未达成共识。为了阐明整个超家族电荷选择性的分子基础，我们对来自脊椎动物、无脊椎动物和细菌的六种不同的阳离子选择性和阴离子选择性同源物进行了广泛的诱变和电生理记录。我们提供了令人信服的证据，证明无论是面向孔道还是埋藏的离子化侧链而非主链原子起着关键作用，并提出了一种机制，据此不仅它们的电荷符号而且它们的构象决定了电荷选择性。涉及孔道胞内端不可电离残基的插入、缺失和残基间突变似乎通过改变M2 α螺旋第一圈中离子化侧链的旋转异构体偏好来影响电荷选择性。我们还发现，在中和M2第一圈中的带电残基后，电荷选择性的控制就移交给了修饰孔道的许多其他离子化侧链。这解释了一个长期存在的谜题，即为什么胞内口谷氨酸的中和在不同的阳离子选择性pLGICs中对电荷选择性的影响程度明显不同。
