# A Genetically Encoded Biosensor for Characterizing Transport and Metabolism of Glutarate

## TL;DR

A new biosensor called Glusor is developed to detect glutarate levels in body fluids and cells, aiding in diagnosing metabolic disorders and studying glutarate metabolism.

## Contribution

The novel contribution is the development of Glusor, a genetically encoded biosensor for glutarate detection with applications in both clinical and metabolic research.

## Key findings

- Glusor accurately quantifies glutarate in human fluids and bacterial fermentation.
- Transporters KgtP and YnfM are identified for glutarate uptake and efflux in E. coli.
- Glusor reveals glutarate catabolism in bacteria and HEK293FT cells and enables spatial analysis.

## Abstract

Glutarate is a platform chemical with diversified applications. It is also an endogenous metabolite involved in various physiological processes. Deficiency in glutaryl‐CoA dehydrogenase (GcdH) for glutarate catabolism induces the inherited metabolic disorder glutaric aciduria. In this study, a genetically encoded glutarate fluorescent biosensor Glusor is constructed and optimized based on transcriptional regulator CsiR and circularly permuted yellow fluorescent protein. Glusor can quantify glutarate in human body fluids and bacteria fermentation broth with good accuracy and precision, supporting the convenient diagnosis of glutaric aciduria and glutarate production monitoring. Then, the glutarate transport is characterized independent of radioactive substrate by using Glusor expressed in Escherichia coli. The functions of transporters KgtP and YnfM in the uptake and efflux of glutarate in E. coli are identified. Glusor is also used to reveal the catabolism of glutarate in bacteria and HEK293FT cells. The role of glutarate hydroxylase in glutarate catabolism of E. coli is identified through Glusor‐supported in situ glutarate detection. Spatially resolved in vivo analysis of glutarate in HEK293FT cells is realized and GcdH inhibition and hypoxia‐induced glutarate accumulation are elucidated by using Glusor. Overall, Glusor is a versatile tool for the detection of glutarate both in vitro and in vivo.

Here, a genetically encoded glutarate biosensor, Glusor, is developed based on transcriptional regulator CsiR. Glusor can quantify glutarate concentrations with good accuracy and precision. Then, the role of KgtP and YnfM are identified and characterized by using Glusor. Glusor also allows glutarate spatiotemporal resolution in live cells, facilitating the elucidation of GcdH inhibition and hypoxia‐induced glutarate accumulation.