Temperature-Dependent Rotation of Protonated Methyl Groups in Otherwise Deuterated Proteins Modulates DEER Distance...

Area of Science:

• Biophysics
• Structural Biology
• Protein Dynamics

Background:

• Electron-Electron Double Resonance (DEER) is a powerful technique for measuring distances in biomolecules.
• Understanding protein dynamics and conformational heterogeneity is crucial for deciphering biological function.

Purpose of the Study:

• To investigate the impact of methyl group rotation on temperature-dependent DEER measurements.
• To develop methods for analyzing complex biomolecular conformations using DEER.

Main Methods:

• Utilized site-specific protonated methyl groups in a deuterated protein background.
• Applied temperature-dependent DEER spectroscopy.
• Analyzed dipolar evolution time traces using a three-site jump model.

Main Results:

• Observed temperature-dependent DEER effects correlated with methyl group rotation (leucine or nitroxide-specific).
• Demonstrated that proximity of a single protonated methyl group selectively decreased the relaxation time (T_m) of the nitroxide label.
• Showcased differential decay of distance distribution components based on temperature and evolution time.

Conclusions:

• Temperature-assisted T_m filtering can enhance DEER structural analysis of heterogeneous biomolecular systems.
• This approach is applicable to complex systems, including multimeric protein complexes.
• Methyl group dynamics provide valuable insights into protein structure and function.