5 years ago

Catch and Release: Engineered Allosterically Regulated β-Roll Peptides Enable On/Off Biomolecular Recognition

Catch and Release: Engineered Allosterically Regulated β-Roll Peptides Enable On/Off Biomolecular Recognition
Mark Blenner, Kevin Dooley, Géza Szilvay, Scott Banta, Beyza Bulutoglu
Alternative scaffolds for biomolecular recognition are being developed to overcome some of the limitations associated with immunoglobulin domains. The repeat-in-toxin (RTX) domain is a repeat protein sequence that reversibly adopts the β-roll secondary structure motif specifically upon calcium binding. This conformational change was exploited for controlled biomolecular recognition. Using ribosome display, an RTX peptide library was selected to identify binders to a model protein, lysozyme, exclusively in the folded state of the peptide. Several mutants were identified with low micromolar dissociation constants. After concatenation of the mutants, a 500-fold increase in the overall affinity for lysozyme was achieved leading to a peptide with an apparent dissociation constant of 65 nM. This mutant was immobilized for affinity chromatography experiments, and the on/off nature of the molecular recognition was demonstrated as the target is captured from a mixture in the presence of calcium and is released in the absence of calcium as the RTX peptides lose their β-roll structure. This work presents the design of a new stimulus-responsive scaffold that can be used for environmentally responsive specific molecular recognition and self-assembly.

Publisher URL: http://dx.doi.org/10.1021/acssynbio.7b00089

DOI: 10.1021/acssynbio.7b00089

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