Structure-guided transformation of channelrhodopsin into a light-activated chloride channel.
Using light to silence electrical activity in targeted cells is a major goal of optogenetics. Available optogenetic proteins that directly move ions to achieve silencing are inefficient, pumping only a single ion per photon across the cell membrane rather than allowing many ions per photon to flow through a channel pore. Building on high-resolution crystal-structure analysis, pore vestibule modeling, and structure-guided protein engineering, we designed and characterized a class of channelrhodopsins (originally cation-conducting) converted into chloride-conducting anion channels. These tools enable fast optical inhibition of action potentials and can be engineered to display step-function kinetics for stable inhibition, outlasting light pulses and for orders-of-magnitude-greater light sensitivity of inhibited cells. The resulting family of proteins defines an approach to more physiological, efficient, and sensitive optogenetic inhibition.
Department of Bioengineering, Stanford University, Stanford, CA 94305, USA.
Andre
A
Berndt
Soo Yeun
SY
Lee
Charu
C
Ramakrishnan
Karl
K
Deisseroth
D000200:Action Potentials; D000595:Amino Acid Sequence; D000818:Animals; D056547:CA1 Region, Hippocampal / Q000166:cytology; D056654:CA3 Region, Hippocampal / Q000166:cytology; D018118:Chloride Channels / Q000737:chemistry* / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism*; D057809:HEK293 Cells; D006801:Humans; D008027:Light; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009474:Neurons / Q000502:physiology*; D062308:Optogenetics; D018408:Patch-Clamp Techniques; D015202:Protein Engineering; D051381:Rats; D017207:Rats, Sprague-Dawley; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D012243:Rhodopsin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
D018118:Chloride Channels; D002712:Chlorides; D011993:Recombinant Fusion Proteins; D012243:Rhodopsin