Biomimetic, Peptide-guided Silica Formation by Real-time NMR of Elastin-like and R5 Fusion Peptides - Bimodal Peptide Aggregation Drives Dual-pathway Silicification Mechanisms.

Abstract

Biomimetic silica particles are promising materials for applications in drug delivery, enzyme encapsulation, and environmental technologies due to their intrinsic biocompatibility and eco-friendly nature. Often, these composites are formed via peptide self-assemblies that can scavenge silicate from solution under mild conditions, thus acting as templates for silica coatings. However, the molecular complexities and large sizes of the peptide assemblies, often exceeding megadaltons, pose significant challenges for structural and functional characterization, leaving key mechanistic aspects unresolved and often impeding rational materials design. Here, we aim to help overcome this bottleneck using methyl-detected Nuclear Magnetic Resonance (NMR) spectroscopy. We investigate a variant of the biotechnologically important R5 peptide, which can template the formation of highly defined silica nanoparticles with variable morphologies. In particular, we focus on elastin-like polypeptide (ELP)-R5 fusion constructs, which have recently been suggested as a promising drug delivery platform. The ELP tag allows for spontaneous self-assembly of R5 above a critical lower solution temperature without the need for any additives that trigger peptide condensation. Exploiting strong methyl resonance intensities and the suppression of the influence of spurious proton exchange on resonance line widths, we were able to access the ELP-R5 peptides within these assemblies. We could follow their resonances even throughout the nanoparticle formation event, simultaneously in real-time and at residue resolution. Integrating our NMR approach with scanning-electron microscopy and dynamic light scattering, we find a previously unrecognized bimodal size distribution of peptide aggregates, giving rise to dual silicification pathways with distinct kinetics.