Pless Stephan A, Millen Kat S, Hanek Ariele P, Lynch Joseph W, Lester Henry A, Lummis Sarah C R, Dougherty Dennis A
School of Biomedical Sciences and Queensland Brain Institute, University of Queensland, Brisbane, Queensland 4072, Australia.
J Neurosci. 2008 Oct 22;28(43):10937-42. doi: 10.1523/JNEUROSCI.2540-08.2008.
Cys-loop receptor binding sites characteristically contain many aromatic amino acids. In nicotinic ACh and 5-HT3 receptors, a Trp residue forms a cation-pi interaction with the agonist, whereas in GABA(A) receptors, a Tyr performs this role. The glycine receptor binding site, however, contains predominantly Phe residues. Homology models suggest that two of these Phe side chains, Phe159 and Phe207, and possibly a third, Phe63, are positioned such that they could contribute to a cation-pi interaction with the primary amine of glycine. Here, we test this hypothesis by incorporation of a series of fluorinated Phe derivatives using unnatural amino acid mutagenesis. The data reveal a clear correlation between the glycine EC(50) value and the cation-pi binding ability of the fluorinated Phe derivatives at position 159, but not at positions 207 or 63, indicating a single cation-pi interaction between glycine and Phe159. The data thus provide an anchor point for locating glycine in its binding site, and demonstrate for the first time a cation-pi interaction between Phe and a neurotransmitter.
半胱氨酸环受体结合位点的特征是含有许多芳香族氨基酸。在烟碱型乙酰胆碱受体和5-羟色胺3型受体中，一个色氨酸残基与激动剂形成阳离子-π相互作用，而在γ-氨基丁酸A型受体中，这一作用由酪氨酸完成。然而，甘氨酸受体结合位点主要含有苯丙氨酸残基。同源性模型表明，这些苯丙氨酸侧链中的两个，即苯丙氨酸159和苯丙氨酸207，可能还有第三个苯丙氨酸63，其位置使它们可能与甘氨酸的伯胺形成阳离子-π相互作用。在此，我们通过使用非天然氨基酸诱变掺入一系列氟化苯丙氨酸衍生物来检验这一假设。数据显示，甘氨酸的半数有效浓度（EC50）值与氟化苯丙氨酸衍生物在位置159处的阳离子-π结合能力之间存在明显相关性，但在位置207或63处则不存在，这表明甘氨酸与苯丙氨酸159之间存在单一的阳离子-π相互作用。因此，这些数据为在其结合位点定位甘氨酸提供了一个锚定点，并首次证明了苯丙氨酸与神经递质之间的阳离子-π相互作用。
