Hydrogels, noted for their biomimetic properties, are the main materials used in biomedical applications such as drug delivery and stem cell therapy. Traditional hydrogels made up of either synthetic polymers or natural biomolecules often serve as passive scaffolds for molecular or cellular species. These features render traditional hydrogels unable to fully recapitulate biological dynamics. It is, therefore, increasingly important to design stimuli-responsive, dynamic hydrogels that can accommodate or mimic the complexity of biological systems. The properties of photoresponsive hydrogels can be controlled using light with great spatiotemporal precision. Although a great number of photoreceptors proteins have been discovered and engineered in the past few decades, which has furt...[ Read more ]

Hydrogels, noted for their biomimetic properties, are the main materials used in biomedical applications such as drug delivery and stem cell therapy. Traditional hydrogels made up of either synthetic polymers or natural biomolecules often serve as passive scaffolds for molecular or cellular species. These features render traditional hydrogels unable to fully recapitulate biological dynamics. It is, therefore, increasingly important to design stimuli-responsive, dynamic hydrogels that can accommodate or mimic the complexity of biological systems. The properties of photoresponsive hydrogels can be controlled using light with great spatiotemporal precision. Although a great number of photoreceptors proteins have been discovered and engineered in the past few decades, which has further led to the emergence of powerful optogenetic tools for numerous biomedical research and applications, most photoresponsive materials still rely on synthetic chromophores as light-sensing moieties, leaving the diverse protein-based photoreceptors untapped.

In this thesis, we developed a new category of protein-based photoresponsive hydrogels by combining the use of recombinant DNA technology, a robust protein assembly strategy known as SpyTag/SpyCatcher chemistry and the newly discovered protein photoreceptor CarH. In Chapter 2, we show the synthesis of the entirely protein-based photoresponsive hydrogels by covalently polymerizing the adenosylcobalamin (AdoB₁₂)-dependent photoreceptor C-terminal adenosylcobalamin binding domain (CarH_C) proteins using genetically encoded SpyTag-SpyCatcher chemistry under mild physiological conditions. Also shown is the resulting hydrogel that exhibited a rapid gel-sol transition on the light exposure. In Chapter 3, we demonstrate that the hydrogel composed of physically self-assembled CarH_C polymers enabled the light-controlled facile release/recovery of human mesenchymal stem cells (hMSCs), 3T3 fibroblasts and PC12 cells from 3D cultures while maintaining their viability. This suggests its great potential as a powerful platform for 3D cell culturing and release. In Chapter 4, a designed covalently cross-linked CarH_C hydrogel can be seen to encapsulate and release bulky globular proteins, such as mCherry, in a light-dependent manner, showing the suitability of using our material for optically controlled release. In Chapter 5, we further explore the feasibility of using other photoresponsive proteins to create hydrogels with different light responsiveness in the hope of gaining insight on some general principles of designing photoresponsive protein materials.

In summary, the direct assembly of stimuli-responsive proteins into hydrogels represents a versatile strategy to design dynamically tunable materials, which may open up new opportunities for materials biology.