Hybrid Two-Component Sensors for Identification of Bacterial Chemoreceptor Function

Area of Science:

• Microbiology
• Biochemistry
• Molecular Biology

Background:

• Soil bacteria utilize complex sensory systems to adapt to environmental changes.
• Two-component systems and chemotaxis pathways are crucial for bacterial signal transduction and environmental response.
• Identifying the specific ligands (chemoeffectors) for bacterial chemoreceptors remains a significant challenge.

Purpose of the Study:

• To develop a simplified method for identifying chemoeffectors sensed by bacterial chemoreceptors.
• To construct and validate hybrid sensor proteins for detecting specific chemical attractants.
• To enable rapid and sensitive screening of potential chemoeffectors for bacterial chemoreceptors.

Main Methods:

• Constructed hybrid sensor proteins by fusing sensor domains of *Pseudomonas putida* chemoreceptors with *Escherichia coli* nitrate sensor signaling domains.
• Utilized a *E. coli* reporter strain with a nitrate-responsive promoter fused to *lacZ* to monitor responses via β-galactosidase assays.
• Performed chemotaxis assays in the native *P. putida* strain to validate findings from the reporter system.

Main Results:

• Hybrid receptors derived from PcaY, McfR, and NahY specifically detected known attractants for aromatic acids, TCA cycle intermediates, and naphthalene.
• β-galactosidase activities in the *E. coli* reporter system correlated well with chemotaxis assay results.
• Identified six novel receptors that detect propionate, expanding the known ligand repertoire for bacterial chemoreceptors.

Conclusions:

• Hybrid sensor proteins coupled with an *E. coli* reporter system provide a rapid, sensitive, and effective method for identifying bacterial chemoeffectors.
• This approach facilitates functional characterization of numerous uncharacterized chemoreceptors in bacterial genomes.
• The developed system holds potential for creating sensitive biosensors for diverse environmental applications, including biodegradation and host-microbe interactions.