FG 1279/2: Channelrhodopsin Colour Tuning (SP 08)

During the past 3 years we have constructed color-tuned high-efficiency channelrhodopsins (ChRs), based on chimeras of Chlamydomonas Channel?rhodopsin-1 (C1) and Volvox Channel?rhodopsin-1 (V1). Several of these variants (C1V1) show superb expression and plasma membrane integration, resulting in 3-fold larger photocurrents in HEK-cells compared to channelrhodopsin-2. Further molecular engineering of C1V1 gave rise to variants with absorption maxima ranging from 526 to 545 nm, matching well with maxima of channelrhodopsin-2 derivatives ranging from 461 to 492 nm. In the project we are aiming at designing new ChRs with absorption maxima covering the whole visible range from 400 to beyond 620 nm. The design will be based on ChRs with most red or blue shifted spetra, on theoretical calculations, homology modelling, and the employment of retinal analogs (synthesized by Dirk Trauners group). Functionality of the new proteins will be studied using electrophysiology and spectroscopy. The new proteins will be tested in C.elegans (with A.Gottschlk) and Zebrafish (with S.Ryu), in hippocampal neurons and living rodents. We also will engineer ChRs with OH-retinal chromophores to be used in Drosophila. Using retinal analogs and enginered ChRs we will test for fluorescence and voltage dependence in order to make a voltage sensitive fluorescent dyes that are able to optically monitor action potentials in neuronal networks.