Channelrhodopsin-2 Function is Modulated by Residual Hydrophobic Mismatch with the Surrounding Lipid Environment

Abstract

Channelrhodopsin-2 (ChR2) is a light-gated ion channel that conducts cations of multiple valencies down the electrochemical gradient. This light-gated property has made ChR2 a popular tool in the field of optogenetics, allowing for the spatial and temporal control of excitable cells with light. A central aspect of protein function is the interaction with the surrounding lipid environment. To further explore these membrane-protein interactions, we demonstrate the role of residual hydrophobic mismatch (RHM) as a mechanistically important component of ChR2 function. We combined computational and functional experiments to understand how RHM between the lipid environment and ChR2 alters the structural and biophysical properties of the channel. Analysis of our results revealed significant RHM at the intracellular/lipid interface of ChR2 from a triad of residues. The resulting energy penalty is substantial and can be lowered via mutagenesis to evaluate the functional effects of this change in lipid-protein interaction energy. The experimental measurement of channel stability, conductance and selectivity resulting from the reduction of the RHM energy penalty showed changes in progressive H+ permeability, kinetics and open-state stability, suggesting how the modulation of ChR2 by the surrounding lipid membrane can play an important biological role and contribute to the design of targeted optogenetic constructs for specific cell types.

Channelrhodopsin C1C2: Photocycle kinetics and interactions near the central gate

“…Expression of functional ChRs has been demonstrated in a variety of different cell types including human neurons, human embryonic kidney cells, Drosophila flies, Caenorhabditis elegans nematodes, and others, all with vastly different membrane compositions. A recent study by our lab found that ChR2 interacts with the surrounding lipid environment and modulates some of its functional characteristics (36). However, although the absolute values of ChR kinetic parameters may differ between expression systems, the trends of the changes observed upon mutation relative to the wild-type channel are expected to remain consistent across cell types, as is the case for many popular ChR variants (12,37).…”