Coupled molecular dynamics mediate long- and short-range epistasis between mutations that affect stability and...

Area of Science:

• Biochemistry
• Protein Engineering
• Computational Biology

Background:

• Improving protein stability and aggregation kinetics often requires multiple mutations.
• Mutations can interact non-additively (epistasis), hindering protein engineering efforts.
• Understanding epistasis is crucial for rational protein design.

Purpose of the Study:

• To investigate the role of protein dynamics in mediating epistatic interactions between mutations.
• To explore epistasis in *Escherichia coli* transketolase (TK) variants.
• To determine if distant mutations can exhibit epistatic effects.

Main Methods:

• Constructed and analyzed specific variants of *Escherichia coli* transketolase (TK).
• Assessed epistasis using measures of protein stability: free-energy barrier to inactivation (∆∆G‡), thermal transition midpoint (Tm), and aggregation onset temperature (Tagg).
• Employed molecular-dynamics simulations and pairwise cross-correlation analysis to study protein dynamics.

Main Results:

• Observed nonadditive epistasis between both neighboring and distant mutations in TK.
• Found that epistatic effects differed across various stability measures (kinetic vs. thermodynamic).
• Demonstrated that mutations can alter dynamics in distant structural regions, mediating epistasis.

Conclusions:

• Protein dynamics play a key role in mediating epistatic interactions, even between distant mutations.
• Kinetic and thermodynamic stability are not always governed by the same structural features.
• Exploiting mutation-induced dynamic changes offers a promising avenue for future protein engineering strategies.