Chemotactic Signaling by Single-Chain Chemoreceptors

Area of Science:

• Microbiology
• Molecular Biology
• Biochemistry

Background:

• Bacterial chemotaxis relies on methyl-accepting chemotaxis proteins (MCPs) forming signaling networks.
• MCPs form homodimers within trimers, enabling precise environmental gradient detection.
• Subunit interactions within MCPs are crucial for signal transduction.

Purpose of the Study:

• To create and test single-chain Tsr receptor molecules for independent subunit manipulation.
• To investigate the impact of linker length on Tsr receptor function and signaling.
• To understand subunit interactions in engineered single-chain chemoreceptors.

Main Methods:

• Construction and characterization of fused genes for single-chain Tsr receptors (Tsr~Tsr).
• Soft agar assays to assess chemotaxis responses.
• Förster resonance energy transfer (FRET)-based in vivo kinase assays to measure CheA activity.

Main Results:

• Single-chain Tsr~Tsr molecules supported serine sensing and chemotaxis.
• Short linkers (≤9 residues) resulted in a constitutively active kinase state.
• Longer linkers (≥22 residues) allowed for near-normal receptor function.
• Evidence of subunit swapping interactions in engineered receptors.

Conclusions:

• Single-chain chemoreceptors are viable tools for studying receptor function.
• Linker length is a critical determinant of single-chain receptor activity.
• Further studies must account for subunit swapping in single-chain receptor applications.