THESIS
2019
xii, 139 pages : illustrations (some color) ; 30 cm
Abstract
Engineered protein hydrogels have emerged as important materials for biomedical applications
thanks to their precisely controlled structure and function and their resemblance towards native
extracellular matrices. Meanwhile, the design of proteins with nonlinear topologies has arisen
as a new branch of protein engineering, yet of which significant applications remain to be seen.
This thesis reports the creation of new hydrogel materials using the protein building blocks
with uncommon topological architectures.
In Chapter 1, the development and applications of protein topology engineering and
protein-based hydrogels are briefly introduced.
In Chapter 2, 3 and 4, we demonstrate the cellular synthesis of a 4-arm star protein,
(SpyCatcher)
4GFP, enabled by spontaneous split-GFP reco...[
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Engineered protein hydrogels have emerged as important materials for biomedical applications
thanks to their precisely controlled structure and function and their resemblance towards native
extracellular matrices. Meanwhile, the design of proteins with nonlinear topologies has arisen
as a new branch of protein engineering, yet of which significant applications remain to be seen.
This thesis reports the creation of new hydrogel materials using the protein building blocks
with uncommon topological architectures.
In Chapter 1, the development and applications of protein topology engineering and
protein-based hydrogels are briefly introduced.
In Chapter 2, 3 and 4, we demonstrate the cellular synthesis of a 4-arm star protein,
(SpyCatcher)
4GFP, enabled by spontaneous split-GFP reconstitution. In combination with the
covalent bond-forming SpyTag/SpyCatcher chemistry, this tetra-functional protein serves as a
modular building block for creating fluorescent protein networks with varied mechanics, well-suited
for cell encapsulation. Conjugating (SpyCatcher)
4GFP with a SpyTagged photoreceptor
protein CarH
C leads to another 4-arm star protein, (CarHC)
4GFP, which undergoes rapid solgel
and gel-sol transitions in response to AdoB
12 and light, respectively. These chemo- and
photo-induced phase transitions enable encapsulation and controlled release of protein
molecules like fluorescent protein mCherry and biofilm-degrading glycosyl hydrolase PslG, a
potential agent for combatting those multidrug-resistant bacterial species such as Pseudomonas
aeruginosa involved in chronic infections.
In Chapter 5, we illustrate the creation of a series of dynamically tunable molecular
networks through the combined use of SpyTag/SpyCatcher chemistry and physically entangled
p53 dimerization domains (Xs). The resulting networks share similar chemical composition
but differ significantly in their viscoelasticity. These materials exhibit excellent compatibility
towards encapsulated fibroblasts and stem cells. Besides, the impact of relaxation rates on
biological processes, like cell morphology developments and bone regenerations are discussed.
Together, this thesis reports the creation of entirely recombinant protein-based
hydrogels using uncommon protein architectures, e.g. four-arm star-like molecules and
physically interlocked structures, of which the applications such as protein release and cell
encapsulation are demonstrated.
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