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ABSTRACT

The primary goal of optogenetics is the light‐controlled noninvasive and specific manipulation of various cellular processes. Herein, we present a hybrid strategy for targeted protein engineering combining computational techniques with electrophysiological and UV/visible spectroscopic experiments. We validated our concept for channelrhodopsin‐2 and applied it to modify the less‐well‐studied vertebrate opsin melanopsin. Melanopsin is a promising optogenetic tool that functions as a selective molecular light switch for G protein‐coupled receptor pathways. Thus, we constructed a model of the melanopsin Gq protein complex and predicted an absorption maximum shift of the Y211F variant. This variant displays a narrow blue‐shifted action spectrum and twofold faster deactivation kinetics compared to wild‐type melanopsin on G protein‐coupled inward rectifying K+ (GIRK) channels in HEK293 cells. Furthermore, we verified the in vivo activity and optogenetic potential for the variant in mice. Thus, we propose that our developed concept will be generally applicable to designing optogenetic tools.

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