Volume and compressibility differences between protein conformations revealed by high-pressure NMR

Area of Science:

• Biophysics
• Structural Biology
• Protein Dynamics

Background:

• Proteins undergo conformational changes essential for their biological functions.
• Manipulating protein conformational equilibria for study is challenging.
• Applying hydrostatic pressure can alter conformational populations by favoring states with lower volumes.

Purpose of the Study:

• To investigate the conformational interconversion of ARNT PAS-B Y456T using high-pressure techniques.
• To quantitatively test hypotheses regarding the volume and compressibility of the slipped conformation and the nature of the intermediate state.

Main Methods:

• High-pressure biomolecular Nuclear Magnetic Resonance (NMR) spectroscopy.
• Steady-state pressure measurements to assess conformational populations.
• Time-resolved pressure-jump experiments to study interconversion kinetics.

Main Results:

• Quantitative data were obtained for the ARNT PAS-B Y456T conformational switch.
• Pressure-dependent changes in conformer populations and interconversion rates were observed.
• The results support the hypotheses that the slipped conformation is smaller and less compressible, and that interconversion proceeds via an unfolded intermediate.

Conclusions:

• High-pressure NMR is a powerful tool for studying protein conformational switches.
• The study provides unique insights into the mechanism of ARNT PAS-B Y456T conformational change.
• These methods can be broadly applied to other protein systems exhibiting conformational dynamics.