posted on 2024-03-13, 21:07authored byWeiyan Zhou, Conor L. O’Neill, Tianben Ding, Oumeng Zhang, Jai S. Rudra, Matthew D. Lew
Synthetic peptides that self-assemble
into cross-β
fibrils
are versatile building blocks for engineered biomaterials due to their
modularity and biocompatibility, but their structural and morphological
similarities to amyloid species have been a long-standing concern
for their translation. Further, their polymorphs are difficult to
characterize by using spectroscopic and imaging techniques that rely
on ensemble averaging to achieve high resolution. Here, we utilize
Nile red (NR), an amyloidophilic fluorogenic probe, and single-molecule
orientation–localization microscopy (SMOLM) to characterize
fibrils formed by the designed amphipathic enantiomers KFE8L and KFE8D and the pathological amyloid-beta peptide Aβ42.
Importantly, NR SMOLM reveals the helical (bilayer) ribbon structure
of both KFE8 and Aβ42 and quantifies the precise tilt of the
fibrils’ inner and outer backbones in relevant buffer conditions
without the need for covalent labeling or sequence mutations. SMOLM
also distinguishes polymorphic branched and curved morphologies of
KFE8, whose backbones exhibit much more heterogeneity than those of
typical straight fibrils. Thus, SMOLM is a powerful tool to interrogate
the structural differences and polymorphism between engineered and
pathological cross-β-rich fibrils.