Targeting metastable coiled-coil domains by computational design

Area of Science:

• Biochemistry
• Structural Biology
• Computational Biology

Background:

• Eukaryotic genomes encode many partially unfolded proteins with transient interactions crucial for cellular homeostasis.
• Current protein design methods struggle with these intrinsically disordered or metastable protein domains.
• Understanding these transient binding events is key to cellular function and disease.

Purpose of the Study:

• To develop and validate a computational design strategy for creating specific peptide inhibitors targeting metastable protein domains.
• To address the limitation of current protein design methods that primarily focus on stable protein structures.
• To create peptide inhibitors for the C-terminal coiled-coil domain of yeast Cdc12p.

Main Methods:

• Implemented a computational design strategy alternating between fixed backbone sequence search and structural interface optimization.
• Applied the method to design peptide inhibitors for the metastable coiled-coil domain of yeast Cdc12p.
• Validated specific binding using biophysical techniques like circular dichroism and equilibrium ultracentrifugation.

Main Results:

• Successfully designed and synthesized specific peptide inhibitors targeting the Cdc12p coiled-coil domain.
• Demonstrated specific binding of the designed peptides to the target protein domain.
• Validated the computational approach for designing ligands to intrinsically unstable protein structures.

Conclusions:

• The developed computational method enables the design of specific peptide ligands for protein domains lacking stable structures.
• This work validates computational approaches for targeting transient protein-protein interactions.
• The designed inhibitors provide tools for future in vivo functional analysis of Cdc12p coiled-coil function.