Development of Light-Gated Ion Channels With Red-Shifted Action Spectra

这个项目的目的是对自然产生的“盲目”受体重新编程，使它们对光变得敏感。利用化学合成和蛋白质工程的结合，我们已经能够将亲离子受体，即离子通道，转化为光感受器。这些可以用来控制生物体内的神经活动和行为。在这里，我们建议使用X射线结晶学(与I.Schlichting合作)和分子动力学模拟在原子水平上研究我们最成功的系统-光致离子型谷氨酸受体(LiGluR)。LiGluR以红藻氨酸受体(IGluR6)为基础，其胞外配体结合域与终止于谷氨酸配体的偶氮苯相连。这种分子的扩展形式和弯曲形式之间的光化学转换改变了配体的有效浓度，允许对受体进行光学控制。我们认为，这个概念是非常普遍的，也可以很容易地应用于代谢性谷氨酸受体。与A.Gottschalk一起，我们还将把它应用于五聚体配体门控离子通道，例如线虫中发现的某些烟碱型乙酰胆碱受体。随后将与J.Heberle合作对该系统进行详细的生物物理研究。我们的合作者S.Ryu和A.Gottschalk将使用优化的光选通谷氨酸和乙酰胆碱受体来解决斑马鱼和线虫(线虫)的神经生物学问题。

The aim of this project is to reprogram naturally occurring “blind” receptors in such a way that they become sensitive toward light. Using a combination of chemical synthesis and protein engineering, we have been able to convert ionotropic receptors, i.e. ion channels, into photoreceptors. These can be used to control neural activity and behavior in living organisms. Herein, we propose to investigate our most successful system, the light-actuated ionotropic glutamate receptor (LiGluR), at the atomic level using X-ray crystallography (in collaboration with I. Schlichting) and with molecular dynamics simulations. LiGluR is based on kainate receptors (iGluR6), whose extracellular ligand-binding domain is conjugated to an azobenzene that terminates in a glutamate ligand. Photochemical switching between the extended and bent form of this molecule changes the effective concentration of the ligand, allowing for optical control of the receptor. We believe that this concept is very general and could be easily applied to metabotropic glutamate receptors as well. Together with A. Gottschalk, we will also apply it to pentameric ligand gated ion channels, such as certain nicotinic acetylcholine receptors found in C. elegans. Detailed biophysical investigations of this system will be subsequently carried out in collaboration with J. Heberle. The optimized lightgated glutamate and acetylcholine receptors will be used by our collaborators S. Ryu and A. Gottschalk to address neurobiological questions in zebrafish and nematodes (C. elegans).