Controlled supramolecular assembly of peptides via chemical reactions
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Abstract
Peptide-based bioresponsive systems have gained significant attention in supramolecular chemistry and biomaterials. These systems have enabled the development of synthetic platforms within living cells, providing valuable insights into how structural formation influences cellular processes. This thesis investigates the design, synthesis, and characterization of different stimulus-responsive peptide systems for the in situ formation of intracellular nanostructures, and examines their consequent effects on cellular processes.
In the first chapter, a platinum (II)-containing tripeptide was engineered to aggregate into intracellular fibrillar nanostructures, disrupting metabolic functions such as aerobic glycolysis and oxidative phosphorylation, consequently instigating apoptosis systematically. This system offers insights into novel building blocks for the construction of peptide-assembling platforms and elucidates the biological mechanisms of action and biochemical profiles associated with nanostructure formation, which implicates system-level effects, such as the regulation of energy and redox homeostasis.
In the second chapter, the development of pH/oxidative stress-responsive isopeptide system was described, including the real-time tracking of the assembling processes and illustration of formed nanostructures, hollow spherical nanofibers, in epithelial cells (MDA-MB 231 cells). Phasor-fluorescence lifetime imaging (FLIM) was utilized to map the transformation form monomers to supramolecular assemblies and correlative light-electron microscopy/tomography (CLEM) revealed formation of nanofibers and their fusion with endosomes to finally form hollow fiber clusters. Spatiotemporal splicing of the assembly events shows time-correlated metabolic dysfunction. This study paves the way to understand and visualize the supramolecular processes of nanostructure formation in biology to ultimately address aggregation-based dysfunction in diseases.
Finally, the construction of two amphiphilic pro-assembling peptide sequences protected by a visible light-sensitive cage group was presents. The peptides underwent a cascade of visible light-induced molecular and supramolecular transformations to form nanofibers in living cells. The irradiation with light enable full control over the reaction cascade where the monomer generation and concentration in turn regulates the assembly kinetics. Phasor-FLIM traced the formation of various assembly states in cells and revealed subsequent out-of-equilibrium dynamics associated with monomer activation and consumption. This study facilitates precise control over supramolecular events at discrete time points, and the new imaging technologies offer deeper insights into the dynamic assembly processes with native cells.
In summary, this thesis introduces a range of methodologies for controlling and imaging intracellular peptide assembly processes and their associated cellular effects. These findings provide valuable insights into supramolecular assembly mechanisms within complex cellular environments and the role of aggregation-based dysfunction in disease.