A genetically encoded ratiometric indicator for tryptophan

Published in Cell Discovery, 2023

Rongkun Tao^†,✉, Kui Wang^†, Tian-lun Chen^†, Xin-xin Zhang, Jian-bin Cao, Wen-quan Zhao, Jiu-lin Du^✉ & Yu Mu^✉

Abstract

In this study, we fused a circularly permuted superfolder YFP into every insertion site of the flexible loop of bacterial tryptophan repressor protein TrpR, where has a large conformational change upon tryptophan binding. We then performed linker truncations and screened a biosensor with ~1.5-fold fluorescence responses to tryptophan. Semi-rational design was conducted by focusing on the residues close to the chromophore or linkers, yielding a mutant with ~6-fold response. The GRIT sensor (Genetically encoded Ratiometric Indicator for Tryptophan) was obtained after screening ~8000 mutants in the linker saturated mutagenesis library. We also engineered a GRIT control sensor (GRITOL) without responses to tryptophan.

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