Effects of pH on proteins: predictions for ensemble and single-molecule pulling experiments

Area of Science:

• Biophysics
• Computational Biology
• Protein Chemistry

Background:

• Protein conformations are sensitive to environmental factors like pH and mechanical forces.
• Predicting changes in protein thermodynamic states requires quantitative models.
• Understanding pH effects on protein properties is crucial for interpreting experimental data.

Purpose of the Study:

• To present a framework, the Molecular Transfer Model (MTM), for predicting protein properties under varying solution conditions, specifically focusing on pH effects.
• To validate the MTM's accuracy in predicting native-state stability as a function of pH for specific proteins.
• To investigate the combined effects of mechanical force and pH on protein unfolding pathways.

Main Methods:

• Utilized the Molecular Transfer Model (MTM) incorporating molecular simulations and partition functions.
• Integrated experimentally measured pK(a) values to calculate free energy changes due to pH variations.
• Applied the MTM to predict protein responses to constant mechanical force across different pH levels.

Main Results:

• Accurate prediction of native-state stability as a function of pH for chymotrypsin inhibitor 2 (CI2) and protein G.
• Distinct phase diagrams for CI2 and protein G under varying force and pH, highlighting pH-dependent stability.
• CI2 exhibits intermediate unfolding under force at certain pH values, while protein G shows single-step, force-independent unfolding, indicating mechanical brittleness.

Conclusions:

• The MTM provides a robust framework for predicting protein behavior under diverse solution conditions, including combined force and pH effects.
• The model accurately captures pH-dependent stability and force-induced unfolding mechanisms.
• MTM facilitates the prediction of outcomes for both ensemble and single-molecule protein experiments.