Hydrogen exchange differences between chemoreceptor signaling complexes localize to functionally important...

Area of Science:

• Molecular biology
• Biochemistry
• Cellular signaling

Background:

• Bacterial chemotaxis receptors mediate transmembrane signaling crucial for cell survival.
• Ligand binding induces conformational changes, but signal propagation through the cytoplasmic domain remains poorly understood.
• Previous studies proposed dynamic changes in subdomains, yet direct measurement in functional complexes was challenging.

Purpose of the Study:

• To investigate signaling-associated dynamics within functional bacterial chemotaxis receptor complexes.
• To elucidate how these dynamics control the activity of the associated kinase CheA.
• To map dynamic changes to specific receptor subdomains involved in excitation and adaptation.

Main Methods:

• Combined hydrogen exchange mass spectrometry (HX-MS) with vesicle template assembly.
• Assembled functional complexes of the receptor cytoplasmic domain with CheA and CheW.
• Measured hydrogen exchange rates to assess protein subdomain stability and dynamics.

Main Results:

• Identified significant signaling-associated changes in hydrogen exchange within the receptor cytoplasmic domain.
• Localized these dynamic changes to key regions involved in excitation and adaptation responses.
• Observed slower hydrogen exchange in the methylation subdomain and protected exchange in the signaling subdomain in the kinase-activating state.

Conclusions:

• The study provides the first measurements of protein subdomain stability within functional signaling complexes.
• Demonstrates that hydrogen exchange mass spectrometry is a powerful tool for studying protein dynamics in multiprotein complexes.
• Reveals distinct dynamic changes in methylation and signaling subdomains correlating with kinase activation, supporting models of signal propagation.