Leucine Motifs Stabilize Residual Helical Structure in Disordered Proteins

Area of Science:

• Protein structure and dynamics
• Biochemistry
• Molecular biology

Background:

• Intrinsically disordered proteins (IDPs) often contain regions that adopt α-helical structures upon binding to targets, known as helical binding motifs (HBMs).
• Residual structure in the unbound state of HBMs can influence binding affinity and kinetics.

Purpose of the Study:

• To investigate the mechanisms governing the formation of residual helical structure in HBMs.
• To understand the role of specific amino acids in stabilizing helical structures and facilitating target binding.

Main Methods:

• Assembled a dataset of experimental helix contents for 65 peptides containing HBMs that fold-upon-binding.
• Analyzed the correlation between residual and target-bound helix contents.
• Investigated the contribution of specific amino acids, particularly leucine, to helix stabilization and binding through sequence analysis and experimental substitutions.

Main Results:

• The average residual helicity of HBMs is 17%, increasing to 60% upon target binding.
• While helix content changes, the relative positions of helical elements show strong overlap between unbound and bound states.
• HBMs are enriched in helix-promoting amino acids, with leucine residues playing a major role in stabilizing helical structure and mediating target binding.
• Substitution of leucine motifs with other hydrophobic residues (valine, isoleucine) reduced residual helicity, confirming leucine's unique stabilizing role.

Conclusions:

• Leucine residues and leucine motifs are critical for stabilizing residual helical structures in HBMs.
• The unique ability of leucine to stabilize helical elements explains its high occurrence and preference at binding interfaces in HBMs.
• Understanding these mechanisms provides insights into the design and function of protein-binding interactions involving disordered proteins.