An experimentally determined protein folding energy landscape

Area of Science:

• Biophysics
• Structural Biology
• Protein Folding Dynamics

Background:

• Energy landscapes conceptually model protein folding but are challenging to measure experimentally due to complex intermediate states.
• The Drosophila Notch receptor, with its modular ankyrin repeat structure, offers a unique system for studying folding pathways.

Purpose of the Study:

• To experimentally determine a detailed energy landscape for protein folding.
• To investigate the contribution of individual ankyrin repeats and their interactions to the overall folding process.

Main Methods:

• Generation of overlapping protein constructs comprising subsets of the seven ankyrin repeats from the Drosophila Notch receptor.
• Thermodynamic characterization of construct stabilities to derive energy terms associated with each repeat.
• Analysis of energy terms to understand the role of nearest-neighbor interactions in stabilizing protein structure.

Main Results:

• Protein construct stabilities approximated a sum of individual repeat energy terms.
• Each ankyrin repeat was found to be intrinsically unstable but significantly stabilized by interactions with adjacent repeats.
• The derived energy terms defined an equilibrium free energy landscape with an initial free energy barrier.

Conclusions:

• Experimental determination of a detailed protein folding energy landscape is feasible.
• Nearest-neighbor interactions play a crucial role in stabilizing protein structures by overcoming the intrinsic instability of individual domains.
• The identified landscape suggests preferred low-energy pathways for protein folding.