Are protein folding intermediates the evolutionary consequence of functional constraints?

Area of Science:

• Protein folding dynamics and biophysics
• Molecular evolution and protein function
• Computational biology and bioinformatics

Background:

• Proteins can exist in multiple conformational states, including partially structured intermediate states.
• The role of evolutionary processes versus functional constraints in shaping these protein folding landscapes is debated.
• Barstar exhibits complex conformational behavior, making it an ideal model for studying protein folding mechanisms.

Purpose of the Study:

• To investigate the determinants of multistate folding in Barstar.
• To understand whether functional constraints or evolutionary history primarily drives protein folding mechanisms.
• To explore the impact of specific mutations on Barstar's folding landscape and stability.

Main Methods:

• Statistical-mechanical modeling of protein folding.
• Electrostatic potential calculations.
• Molecular dynamics (MD) simulations.
• Multiple-sequence alignment analysis.

Main Results:

• Barstar's multistate folding is driven by evolutionary pressure to maintain binding affinity with Barnase via electrostatic interactions.
• A single point mutation (E76K or E80K) at the binding site stabilizes the native state and simplifies folding to a two-state-like mechanism.
• Functional constraints can override the tendency for minimal frustration, dictating folding pathways.

Conclusions:

• Functional requirements, specifically maintaining protein-protein binding affinity, can singularly dictate a protein's folding mechanism.
• This can lead to frustrated folding landscapes even when minimal frustration is generally expected in natural proteins.
• Evolutionary pressures for function can shape protein folding beyond simply minimizing frustration.