De novo chemo-optogenetics through the rational design of photoresponsive molecules and selection of their artificial protein binding pairs.

Optical manipulation of proteins is central to probing and engineering cellular functions. However, existing optogenetic tools based on natural photoreceptors and chemo-optogenetic tools based on natural protein-ligand pairs are difficult to reconfigure for the desired photochemical and binding properties. Here we introduce a de novo approach for creating chemo-optogenetic tools. Instead of modifying pre-existing ligands, we first design a synthetic photoswitch with defined properties and then use mRNA display to select for artificial protein binders that recognize a specific photoisomer-dependent conformation. This bottom-up framework yields artificial photoswitch-protein binder pairs that enable precise optical control of diverse biological activities in mammalian cells, such as kinase and lipid signalling, G-protein-coupled receptor activation, gene expression and cell differentiation. The regulatory mode (sustained, reversible or repeatable) is readily programmed by adjusting light inputs. This de novo approach provides customizable synthetic photoswitch-protein binder pairs, expanding opportunities for optical protein manipulation in biological and biomedical applications.