Genetically encoded tools for tracking metabolites in live cells

Biosensors enable the in situ measurement of metabolites in living systems over time and space. Fully genetically encoded metabolite biosensors (fGEMBs) use fluorescent proteins (FPs) linked to ligand binding domains (LBDs) to transduce the ligand binding event to a measurable change in the fluorescence behavior of the FP. Because these sensors are genetically encoded, they can be expressed in cells using standard protein expression approaches, and the fluorescence changes are quantified using fluorimetry, fluorescence microscopy, and/or flow cytometry. While there are general sensor design principles to follow, an fGEMB must be engineered for each metabolite based on a particular LBD. This development process can be slow, but there are strategies emerging to increase testing throughput and improve structure-guided design. While genetically-encoded FPs remain popular, there are now numerous chemigenetic and nucleic acid-based metabolite sensors (cGEMBs) that incorporate small molecule fluorophores. De novo design of LBDs is rapidly advancing as well, and the field may soon exhibit a shift away from relying on nature's catalog of LBDs. Despite the engineering challenges, the metabolite biosensor field has expanded significantly in recent years to meet the demand for new and better-performing sensors that visualize metabolites within their cellular environments.