Probing protein ensemble rigidity and hydrogen-deuterium exchange

Area of Science:

• Structural Biology
• Computational Biophysics
• Protein Dynamics

Background:

• Traditional protein rigidity and flexibility analysis uses single static structures.
• Proteins exist as ensembles of native-like conformers due to spontaneous dynamics.
• Existing methods fail to capture the full picture of protein conformational heterogeneity.

Purpose of the Study:

• To develop a novel FIRST-based approach for predicting protein ensemble rigidity and flexibility.
• To integrate ensemble rigidity with solvent accessibility for predicting hydrogen-deuterium exchange (HDX).
• To validate the computational predictions against experimental HDX data.

Main Methods:

• A refined FIRST (floppy inclusion and rigid substructure topography) program was used.
• Averaged hydrogen bonding strengths across the protein ensemble were calculated.
• Ensemble solvent accessibility data was combined with rigidity predictions for HDX modeling.

Main Results:

• The FIRST-ensemble approach accurately predicted sub-structural conformational dynamics.
• Experimental HDX data for Sso AcP closely matched the FIRST-ensemble rigidity predictions.
• Computational predictions of HDX protection and exchange regions agreed well with experimental profiles.

Conclusions:

• Analyzing proteins as dynamic ensembles provides a more accurate representation of their behavior.
• The novel FIRST-ensemble approach successfully predicts protein dynamics and HDX.
• This method offers improved insights into protein structural ensembles and their functional implications.