A Biosensor Strategy for E. coli Based on Ligand-Dependent Stabilization.

Area of Science:

• Synthetic Biology
• Microbial Engineering
• Biochemistry

Background:

• Developing effective biosensors is crucial for environmental monitoring and bioproduct synthesis.
• Converting natural ligand-binding proteins into functional biosensors remains a significant challenge in microbial engineering.

Purpose of the Study:

• To establish a generalizable strategy for engineering ligand-responsive biosensors in Escherichia coli.
• To demonstrate the conversion of ligand-binding proteins into biosensors through directed evolution.

Main Methods:

• Fusion of ligand-binding domains (e.g., lac repressor) with DNA-binding domains and transcription-activating domains.
• Utilizing error-prone PCR mutagenesis and selection to identify functional biosensor variants.
• Engineering ligand specificity and response through targeted mutations.

Main Results:

• A novel biosensor was constructed in E. coli, exhibiting ligand-dependent stabilization of a transcriptional activator.
• A 7-fold increase in E. coli growth rate was achieved, dependent on the inducer isopropyl β-d-1-thiogalactopyranoside (IPTG).
• The strategy was validated by successfully engineering the MphR ligand-binding domain into a functional biosensor.

Conclusions:

• The developed strategy offers a versatile approach for converting diverse ligand-binding proteins into robust biosensors.
• This method facilitates the creation of microbial systems for detecting environmental chemicals and producing valuable bioproducts.
• The findings pave the way for broader applications of engineered biosensors in biotechnology and environmental science.