Remodeling a β-peptide bundle

Natural biopolymers fold with fidelity, burying diverse side chains into well-packed cores and protecting their backbones from solvent. Certain beta-peptide oligomers assemble into bundles of defined octameric stoichiometry that resemble natural proteins in many respects. These beta-peptide bundles are thermostable, fold cooperatively, exchange interior amide N-H protons slowly, exclude hydrophobic dyes, and can be characterized at high resolution using X-ray crystallography - just like many proteins found in nature. But unlike natural proteins, all octameric beta-peptide bundles contain a sequence-uniform hydrophobic core composed of 32 leucine side chains. Here we apply rational design principles, including the Rosetta computational design methodology, to introduce sequence diversity into the bundle core while retaining the characteristic beta-peptide bundle fold. Using circular dichroism spectroscopy and analytical ultracentrifugation, we confirmed the prediction that an octameric bundle still assembles upon a major remodelling of its core: the mutation of sixteen core beta-homo-leucine side chains into sixteen beta-homo-phenylalanine side chains. Nevertheless, the bundle containing a partially beta-homo-phenylalanine core poorly protects interior amide protons from exchange, suggesting molten-globule-like properties. We further improve stability by the incorporation of eight beta-homo-pentafluorophenyalanine side chains, giving an assembly with amide protection factors comparable to prior well-structured bundles. By demonstrating that their cores tolerate significant sequence variation, the beta-peptide bundles reported here represent a starting point for the bottom-up construction of beta-peptide assemblies possessing both structure and sophisticated function.