Rienzo Matthew, Lummis Sarah C R, Dougherty Dennis A
Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125, USA.
Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1GA, UK.
Chem Biol. 2014 Dec 18;21(12):1700-6. doi: 10.1016/j.chembiol.2014.10.019.
The cyanobacterial pentameric ligand-gated ion channel GLIC, a homolog of the Cys-loop receptor superfamily, has provided useful structural and functional information about its eukaryotic counterparts. X-ray diffraction data and site-directed mutagenesis have previously implicated a transmembrane histidine residue (His234) as essential for channel function. Here, we investigated the role of His234 via synthesis and incorporation of histidine analogs and α-hydroxy acids using in vivo nonsense suppression. Receptors were expressed heterologously in Xenopus laevis oocytes, and whole-cell voltage-clamp electrophysiology was used to monitor channel activity. We show that an interhelix hydrogen bond involving His234 is important for stabilization of the open state, and that the shape and basicity of its side chain are highly sensitive to perturbations. In contrast, our data show that two other His residues are not involved in the acid-sensing mechanism.
蓝藻五聚体配体门控离子通道GLIC是半胱氨酸环受体超家族的同源物，它为其真核对应物提供了有用的结构和功能信息。X射线衍射数据和定点诱变先前表明跨膜组氨酸残基（His234）对通道功能至关重要。在此，我们通过使用体内无义抑制合成并掺入组氨酸类似物和α-羟基酸来研究His234的作用。受体在非洲爪蟾卵母细胞中异源表达，并使用全细胞膜片钳电生理学来监测通道活性。我们表明，涉及His234的螺旋间氢键对于开放状态的稳定很重要，并且其侧链的形状和碱性对扰动高度敏感。相比之下，我们的数据表明另外两个His残基不参与酸感应机制。
