Folding Intermediates, Heterogeneous Native Ensembles and Protein Function.

Area of Science:

• Protein folding mechanisms
• Biophysics
• Structural biology

Background:

• Single domain proteins fold through various mechanisms, linking energetics and structure to functional demands.
• Studying folding mechanisms in large proteins is difficult due to experimental and computational limitations in identifying metastable states.

Purpose of the Study:

• To investigate the folding mechanisms and functional imprints in large proteins using a statistical mechanical model.
• To explore the relationship between protein structure, folding energetics, and biological function in systems over 150 residues.

Main Methods:

• Utilized a statistical mechanical model to predict free energy profiles and surfaces for large protein systems (>150 residues).
• Analyzed folding landscapes to identify folding intermediates and heterogeneous native ensembles.

Main Results:

• Large protein systems (>150 residues) display diverse folding mechanisms, including common folding intermediates and varied native ensembles.
• Residues near ligand binding or enzyme active sites show increased propensity for partial unfolding, appearing as intermediates or excited states.
• Observed these functional imprints in various proteins, including SARS-CoV-2 receptor binding domain (RBD) and Mpro protease.

Conclusions:

• It is easier to discern functional imprints on the folding landscapes of larger proteins compared to smaller ones.
• Understanding energetic-entropic features can guide engineering of protein folding, partially structured states, and function.