Characterization of Engineered Channelrhodopsin Variants with Improved Properties and Kinetics
journal contribution
posted on 2023-05-18, 05:54authored byJohn LinJohn Lin, Lin, MZ, Steinbach, P, Tsien, RY
Channelrhodopsin 2 (ChR2), a light-activated nonselective cationic channel from Chlamydomonas reinhardtii, has become a useful tool to excite neurons into which it is transfected. The other ChR from Chlamydomonas, ChR1, has attracted less attention because of its proton-selective permeability. By making chimeras of the transmembrane domains of ChR1 and ChR2, combined with site-directed mutagenesis, we developed a ChR variant, named ChEF, that exhibits significantly less inactivation during persistent light stimulation. ChEF undergoes only 33% inactivation, compared with 77% for ChR2. Point mutation of Ile170 of ChEF to Val (yielding "ChIEF") accelerates the rate of channel closure while retaining reduced inactivation, leading to more consistent responses when stimulated above 25 Hz in both HEK293 cells and cultured hippocampal neurons. In addition, these variants have altered spectral responses, light sensitivity, and channel selectivity. ChEF and ChIEF allow more precise temporal control of depolarization, and can induce action potential trains that more closely resemble natural spiking patterns.
History
Publication title
Biophysical Journal
Volume
96
Pagination
1803-1814
ISSN
0006-3495
Department/School
Tasmanian School of Medicine
Publisher
Biophysical Society
Place of publication
9650 Rockville Pike, Bethesda, USA, Md, 20814-3998