sb7b00089_si_001.pdf (2.97 MB)
Catch and Release: Engineered Allosterically Regulated β‑Roll Peptides Enable On/Off Biomolecular Recognition
journal contribution
posted on 2017-05-18, 00:00 authored by Beyza Bulutoglu, Kevin Dooley, Géza Szilvay, Mark Blenner, Scott BantaAlternative 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.