Orthogonal recognition in dimeric coiled coils via buried polar-group modulation

Area of Science:

• Biochemistry
• Molecular Biology
• Peptide Chemistry

Background:

• Coiled-coil structures are crucial in biological systems.
• Controlling protein-protein interactions, like coiled-coil dimerization, is key for molecular design.
• Introducing buried polar groups offers a novel strategy for precise dimerization control.

Purpose of the Study:

• To design and explore novel buried polar groups for controlling coiled-coil dimerization.
• To develop peptides with tunable stability and orthogonal recognition properties.
• To create a system for forming two distinct heterodimeric coiled coils simultaneously.

Main Methods:

• On-resin guanidinylation for peptide synthesis.
• Preparation of coiled-coil peptides with single core guanidine groups.
• Heterodimeric mixture formation with various binding partners (guanidine, amide, carboxylic acid).
• Biophysical characterization including circular dichroism, ultracentrifugation, and denaturation studies.

Main Results:

• Successfully formed stable coiled coils (Tm ≥ 60°C) using peptides with buried guanidine groups.
• Identified distinct stability trends: asparagine/acid pairs are sensitive to acid chain length, while guanidine/acid pairs are insensitive.
• Demonstrated orthogonal recognition by creating two distinct heterodimeric coiled coils from four peptides.
• Confirmed specificity through the failure of glutamic acid substitutions and extensive biophysical validation.

Conclusions:

• Buried polar groups, particularly guanidine and asparagine, can effectively control coiled-coil dimerization.
• The differential sensitivity of residue pairs allows for the design of orthogonal recognition systems.
• This work provides a powerful tool for constructing complex protein architectures and controlling molecular assembly.