Predicting repeat protein folding kinetics from an experimentally determined folding energy landscape

Area of Science:

• Protein folding dynamics
• Biophysics
• Molecular biology

Background:

• The Notch ankyrin domain is a repeat protein.
• Previous studies characterized its folding using equilibrium measurements and generated a folding energy landscape.

Purpose of the Study:

• To parameterize a kinetic model using the established folding energy landscape.
• To predict and validate the folding kinetics of the Notch ankyrin domain and its variants.

Main Methods:

• Developed a kinetic model based on local transition probabilities derived from the folding energy landscape.
• Utilized experimental data for Notch ankyrin domain folding and sequence variants.

Main Results:

• The landscape-based kinetic model accurately predicted diverse folding kinetics, including monophasic folding and biphasic unfolding.
• The model successfully predicted chevron plot curvature, transient intermediate populations, and folding rates across variants.
• Demonstrated that C-terminal stabilization alters the folding pathway.

Conclusions:

• Protein folding pathways are thermodynamically selected.
• Local repeat stability is a primary determinant of kinetic behavior in the Notch ankyrin domain.
• The folding landscape effectively guides kinetic predictions.