Hierarchical assembly of tryptophan zipper peptides into tissue-mimetic hydrogels

Abstract

In nature, soft functional materials like human tissue are formed through the supramolecular assembly of biological polymers across the nanoscale, microscale, and macroscale. Harnessing the design rules of these naturally occurring hierarchical structures to synthesise our own functional biomimetic materials is an important research goal spanning multiple fields including supramolecular chemistry, materials science, tissue engineering and medicine. Rational design has allowed us to craft self- assembling peptides with structural similarities to natural materials, but recreating their inherently dynamic functional properties remains challenging. In this thesis, I introduce a short self-assembling peptide based on the tryptophan zipper (Trpzip) motif, which shows multiscale hierarchical ordering leading to emergent dynamic properties in the resulting hydrogels. Initially conceived to serve as a simple monomeric model to study β-tertiary structure and folding, I demonstrate that with minor residue substitutions, the Trpzip motif can be repurposed as the minimal structural unit from which nanofibrous, tissue-mimetic hydrogels can be assembled. Using molecular dynamics simulations to navigate the vast sequence space, I identified, synthesized, and validated a Trpzip variant which formed a self-assembling hydrogel under physiological conditions. I then examined the nanoscale and microscale properties of these Trpzip hydrogels through a variety of spectroscopy, microscopy, and neutron scattering techniques. Next, I characterised the mechanics of pure Trpzip hydrogels, finding that they show tunable viscoelasticity, can shear-thin and self-heal like other supramolecular hydrogels yet display unique yield-stress properties, and demonstrate stress-relaxation behavior reminiscent of native tissue. With a variety of other supramolecular and covalent chemistry approaches, I explore ways to further modulate the mechanical and biochemical profile of Trpzip hydrogels. Finally, a range of cellular and organotypic models were integrated into Trpzip hydrogels to demonstrate their potential as synthetic 3D tissue scaffolds. I show pure Trpzip hydrogels support a variety of mammalian cell types including fibroblasts, myoblasts, and induced pluripotent stem cells (iPSCs), and its rheological properties make the hydrogel amenable to clinical and biofabrication applications such as syringe- based cell delivery and 3D bioprinting. Using various human intestinal organoid models, I show Trpzip hydrogels can support complex yet critical biological processes such as self-organization, morphogenesis, and polarization. Altogether, the research presented in this thesis expands the supramolecular peptide design space, introduces a synthetic biomaterial with potential for use in tissue engineering and medicine, and contributes to our growing understanding of the materials chemistry principles central to recapitulating native tissue matrices