Structures of designed armadillo-repeat proteins show propagation of inter-repeat interface effects

Area of Science:

• Protein engineering
• Structural biology
• Biochemistry

Background:

• Armadillo repeats form versatile scaffolds for creating modular peptide-recognition systems.
• Designed armadillo-repeat proteins with specific cap and internal repeat configurations were investigated.

Purpose of the Study:

• To determine the crystal structures of designed armadillo-repeat proteins with varying cap types.
• To compare structural differences influenced by C-cap design and intrinsic repeat properties.
• To assess the impact of these structures on the putative peptide-binding site.

Main Methods:

• X-ray crystallography was used to determine the structures of three designed armadillo-repeat proteins at high resolution (1.8–2.0 Å).
• Comparative structural analysis was performed on variants with second-generation and third-generation C-caps.
• The influence of C-cap interactions and intrinsic repeat design on protein structure was evaluated.

Main Results:

• While N-caps and initial internal repeats (M1-M3) showed high similarity, later repeats (M4-M5) and C-caps exhibited structural variations.
• Long-range effects from the C-cap and inherent repeat design contributed to observed structural differences.
• C-terminal movement negatively affected the regularity of internal repeats crucial for peptide binding.

Conclusions:

• Second-generation C-caps enhance protein stability by improving buried residue packing.
• Structural insights gained are valuable for refining armadillo-repeat-based peptide-recognition systems.
• Understanding these structure-function relationships is key for future protein design advancements.