A designed system for assessing how sequence affects α to β conformational transitions in proteins

The role of amino acid sequence in conformational switching observed in prions and proteins associated with amyloid diseases is not well understood. To study a to P conformational transitions, we designed a series of peptides with structural duality; namely, peptides with sequence features of both an a-helical leucine zipper and a beta-hairpin. The parent peptide, Template-alpha, was designed to be a canonical leucine-zipper motif and was confirmed as such using circular dichroism, spectroscopy and analytical ultracentrifugation. To introduce beta-structure character into the peptide, glutamine residues at sites away from the leucine-zipper dimer interface were replaced by threonine to give Template-alphaT. Unlike the parent peptide, Template-alphaT underwent a heat-inducible switch to beta-structure, which reversibly formed gels containing amyloid-like fibrils. In contrast to certain other natural proteins where destabilization of the native states facilitate transitions to amyloid, destabilization of the leucine-zipper form of Template-alphaT did not promote a transformation. Cross-linking the termini of the peptides compatible with the alternative,beta-hairpin design, however, did promote the change. Furthermore, despite screening various conditions, only the internally cross-linked form of the parent, Template -alpha, peptide formed amyloid-like fibrils. These findings demonstrate that, in addition to general properties of the polypeptide backbone, specific residue placements that favor beta-structure promote amyloid formation.