Localized thermodynamic coupling between hydrogen bonding and microenvironment polarity substantially stabilizes...

Area of Science:

• Biochemistry
• Structural Biology
• Biophysics

Background:

• The role of hydrogen bonding in protein folding remains incompletely understood despite decades of research.
• Extracting precise thermodynamic data for specific interactions from mutagenesis experiments is challenging.
• Hydrogen bond strength is known to be influenced by its surrounding environment.

Purpose of the Study:

• To investigate the hypothesis that hydrogen bond strength is modulated by the polarity of its local microenvironment.
• To quantify the energetic contribution of microenvironmental polarity to hydrogen bond strength in proteins.

Main Methods:

• Utilized double-mutant cycle analysis to assess energetic contributions.
• Employed a combination of amide-to-ester backbone mutagenesis and traditional side chain mutagenesis.
• Compared hydrogen bond strengths in different microenvironments (hydrophobic vs. solvent-exposed).

Main Results:

• Demonstrated that hydrogen bonds can be up to 1.2 kcal mol(-1) stronger when sequestered in hydrophobic surroundings compared to solvent-exposed locations.
• Quantified significant coupling energies between hydrogen bond strength and local polarity.
• Provided thermodynamic data on specific hydrogen bond interactions within a protein context.

Conclusions:

• The polarity of a protein's microenvironment significantly impacts hydrogen bond strength.
• Context dependence of hydrogen bond strengths is a critical factor that must be incorporated into models of protein folding.
• These findings offer new insights into the forces governing protein structure and stability.