Design of a Survival-Fluorescence Cascade Screening Circuit (uFAST) for an Erythritol-Specific BmoR-Based Biosensor

Area of Science:

• Synthetic Biology
• Biotechnology
• Biosensor Development

Background:

• Accurate erythritol quantification is difficult due to its similarity to other sugars.
• Existing methods lack specificity and speed for complex sample analysis.

Purpose of the Study:

• To engineer a highly specific and sensitive biosensor for erythritol detection.
• To improve erythritol production strains using the developed biosensor platform.

Main Methods:

• Developed a survival-fluorescence cascade screening circuit (uFAST) with a σ54-dependent BmoR-based biosensor.
• Utilized SacB-mediated negative and mCherry-mediated positive selection for mutant isolation.
• Optimized the BmoR protein and coupled it with a VioABCE visual module for quantification.

Main Results:

• Achieved a 6.03-fold increase in GFP output and a 2.09-fold improvement in apparent affinity (Km) for the BmoR biosensor.
• The biosensor accurately quantified erythritol in complex matrices, comparable to HPLC results.
• Identified an improved erythritol production strain yielding 3.51 g/L, a 2.88-fold increase in titer.

Conclusions:

• The uFAST circuit provides a robust framework for developing specific biosensors.
• The engineered biosensor is an efficient tool for erythritol detection and strain improvement in biotechnology.
• This platform facilitates rapid screening and optimization of microbial production systems.