Engineering of an artificial light-modulated potassium channel

Caro, Lydia N., Moreau, Christophe J., Estrada-Mondragón, Argel, Ernst, Oliver P. and Vivaudou, Michel (2012) Engineering of an artificial light-modulated potassium channel. PLoS One, 7 8: e43766.1-e43766.9. doi:10.1371/journal.pone.0043766

Author Caro, Lydia N.
Moreau, Christophe J.
Estrada-Mondragón, Argel
Ernst, Oliver P.
Vivaudou, Michel
Title Engineering of an artificial light-modulated potassium channel
Journal name PLoS One   Check publisher's open access policy
ISSN 1932-6203
Publication date 2012
Sub-type Article (original research)
DOI 10.1371/journal.pone.0043766
Open Access Status DOI
Volume 7
Issue 8
Start page e43766.1
End page e43766.9
Total pages 9
Place of publication San Francisco, CA, United States
Publisher Public Library of Science
Language eng
Abstract Ion Channel-Coupled Receptors (ICCRs) are artificial receptor-channel fusion proteins designed to couple ligand binding to channel gating. We previously validated the ICCR concept with various G protein-coupled receptors (GPCRs) fused with the inward rectifying potassium channel Kir6.2. Here we characterize a novel ICCR, consisting of the light activated GPCR, opsin/rhodopsin, fused with Kir6.2. To validate our two-electrode voltage clamp (TEVC) assay for activation of the GPCR, we first co-expressed the apoprotein opsin and the G protein-activated potassium channel Kir3.1F137S (Kir3.1*) in Xenopus oocytes. Opsin can be converted to rhodopsin by incubation with 11-cis retinal and activated by light-induced retinal cis→trans isomerization. Alternatively opsin can be activated by incubation of oocytes with all-trans-retinal. We found that illumination of 11-cis-retinal-incubated oocytes co-expressing opsin and Kir3.1* caused an immediate and long-lasting channel opening. In the absence of 11-cis retinal, all-trans-retinal also opened the channel persistently, although with slower kinetics. We then used the oocyte/TEVC system to test fusion proteins between opsin/rhodopsin and Kir6.2. We demonstrate that a construct with a C-terminally truncated rhodopsin responds to light stimulus independent of G protein. By extending the concept of ICCRs to the light-activatable GPCR rhodopsin we broaden the potential applications of this set of tools.
Keyword Cell membranes
Ion channel gating
Ligand-gated ion channels
Membrane proteins
Potassium channels
Sequence alignment
Xenopus oocytes
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status Non-UQ

Document type: Journal Article
Sub-type: Article (original research)
Collection: Queensland Brain Institute Publications
Version Filter Type
Citation counts: TR Web of Science Citation Count  Cited 6 times in Thomson Reuters Web of Science Article | Citations
Scopus Citation Count Cited 7 times in Scopus Article | Citations
Google Scholar Search Google Scholar
Created: Thu, 04 Sep 2014, 09:19:22 EST by Argel Estrada on behalf of Queensland Brain Institute