1. Affinity Series of Genetically Encoded Förster Resonance Energy-Transfer Sensors for Sucrose
- Author
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Mayuri Sadoine, Mira Reger, Wolf B. Frommer, and Ka Man Wong
- Subjects
Sucrose ,Bioengineering ,02 engineering and technology ,Biosensing Techniques ,01 natural sciences ,chemistry.chemical_compound ,In vivo ,Fluorescence Resonance Energy Transfer ,Animals ,Binding site ,Sugar ,Instrumentation ,Trehalose transport ,Fluid Flow and Transfer Processes ,biology ,Process Chemistry and Technology ,010401 analytical chemistry ,Agrobacterium tumefaciens ,021001 nanoscience & nanotechnology ,biology.organism_classification ,Trehalose ,0104 chemical sciences ,Luminescent Proteins ,Förster resonance energy transfer ,Glucose ,chemistry ,Biochemistry ,0210 nano-technology - Abstract
Genetically encoded fluorescent sugar sensors are valuable tools for the discovery of transporters and for quantitative monitoring of sugar steady-state levels in intact tissues. Genetically encoded Forster resonance energy-transfer sensors for glucose have been designed and optimized extensively, and a full series of affinity mutants is available for in vivo studies. However, to date, only a single improved sucrose sensor FLIPsuc-90μΔ1 with Km for sucrose of ∼90 μM was available. This sucrose sensor was engineered on the basis of an Agrobacterium tumefaciens sugar-binding protein. Here, we took a two-step approach to first improve the dynamic range of the FLIPsuc sensor and then expand the detection range from micro- to millimolar sucrose concentrations by mutating a key residue in the binding site. The resulting series of sucrose sensors may enable investigation of sucrose transporter candidates and comprehensive in vivo analyses of sucrose concentration in plants. Since FLIPsuc-90μ also detects trehalose in animal cells, the new series of sensors will likely be suitable for investigating trehalose transport and monitor trehalose steady-state levels in vivo.
- Published
- 2021