Dynamic Nuclear Polarization Illuminates Key Protein-Lipid Interactions in the Native Bacterial Cell Envelope

Area of Science:

• Structural Biology
• Biophysics
• Microbiology

Background:

• Elucidating protein structure and interactions in native environments is vital for understanding biological processes.
• Nuclear magnetic resonance (NMR) spectroscopy is a powerful tool but often limited by low sensitivity in complex biological systems.
• The outer membrane protein Ail from Yersinia pestis plays a critical role in host invasion.

Purpose of the Study:

• To overcome the sensitivity limitations of conventional NMR for studying proteins in native bacterial environments.
• To investigate the membrane interactions of the Yersinia pestis outer membrane protein Ail using a novel technique.
• To elucidate the role of Ail in host invasion by examining its interactions with the bacterial cell envelope.

Main Methods:

• Application of dynamic nuclear polarization (DNP) to enhance NMR sensitivity.
• Solid-state NMR experiments on the outer membrane protein Ail within native bacterial cell envelopes.
• Analysis of DNP-enhanced NMR spectra to identify protein-lipid interactions.

Main Results:

• DNP-enhanced NMR provided well-resolved spectra of Ail in native bacterial cell envelopes.
• The technique revealed spectral correlations previously undetectable with conventional solid-state NMR.
• Elusive interactions between Ail and the surrounding lipopolysaccharide layer were successfully captured.

Conclusions:

• Dynamic nuclear polarization significantly improves NMR sensitivity for studying membrane proteins in native environments.
• The findings support a model where Ail's extracellular arginine residues remodel the membrane, facilitating host invasion.
• This study highlights DNP-NMR as a valuable method for understanding protein-lipid interactions in bacterial pathogenesis.