Remarkably slow folding of a small protein

Area of Science:

• Protein folding dynamics
• Biophysics of small proteins
• Biochemical kinetics

Background:

• MerP is a 72-amino acid alpha/beta-protein.
• Protein folding is crucial for biological function.
• Understanding folding mechanisms aids in protein engineering and disease research.

Purpose of the Study:

• To investigate the equilibrium denaturation and folding kinetics of the MerP protein.
• To characterize the reversibility and thermodynamic model of MerP unfolding.
• To identify factors contributing to MerP's folding rate.

Main Methods:

• Equilibrium denaturation studies using acid, guanidine hydrochloride, and temperature.
• Analysis of folding kinetics to determine rate constants.
• Characterization of unfolded states and populations.

Main Results:

• MerP denaturation is fully reversible and follows a two-state model.
• A cis-trans equilibrium of a proline peptide bond leads to heterogeneous unfolded states (slow and fast folding populations).
• The fast-folding population of MerP exhibits a rate constant of 1.2 s(-1), indicating unusually slow folding for its size.

Conclusions:

• MerP represents the slowest folding protein of its small size reported to date.
• Proline peptide bond isomerization is a key determinant of MerP's slow folding kinetics.
• The study provides insights into the folding pathways of small proteins and the role of proline isomerization.