A large solvent isotope effect on protein association thermodynamics

Area of Science:

• Biochemistry
• Structural Biology
• Thermodynamics

Background:

• Protein-protein interactions are crucial in biological processes.
• Solvent reorganization is a key factor influencing interaction energetics.
• The biotin repressor dimerizes for DNA binding and transcriptional repression.

Purpose of the Study:

• To quantify the energetic contribution of solvent release to protein dimerization.
• To investigate the role of solvent reorganization in the biotin repressor's homodimerization.
• To validate the interpretation of thermodynamic data through solvent isotope effects.

Main Methods:

• Sedimentation equilibrium measurements were used to determine dimerization thermodynamics.
• Experiments were conducted in both H2O and D2O at 20 °C.
• Temperature dependence studies in D2O were performed to analyze enthalpic and entropic contributions.

Main Results:

• The Gibbs free energy of dimerization for the biotin repressor is -7 kcal/mol at 20 °C.
• Replacing H2O with D2O resulted in a -1.5 kcal/mol solvent isotope effect on dimerization free energy.
• Thermodynamic analysis in D2O showed reduced enthalpic (28 kcal/mol) and entropic (-37 kcal/mol) contributions compared to H2O.

Conclusions:

• The significant solvent isotope effect supports the coupling of solvent release to biotin repressor dimerization.
• Solvent reorganization plays a substantial role in the energetics of this protein-protein interaction.
• These findings provide quantitative experimental evidence for the impact of solvent dynamics on protein complex formation.