From Microscale Network Topology to Macroscale Mechanics of Folded protein Hydrogels

Hydrogels are a unique species of soft matter biomaterial. Generically defined as any hydrophilic polymer network capable of absorbing large volumes of water, their composition and characteristics can be extremely varied. With such an all-encompassing definition, hydrogels represent potentially the most tuneable and adaptable scaffold for biomaterial development in current research, with a myriad of possible applications in both laboratory and clinical settings, ranging from advanced wound dressings to 3-dimensional cell culture models. The majority of hydrogels to date have been built from non-biological polymer lattices, but over the last 15 years research has shifted towards building hydrogels out of natively folded and dynamically active proteins. Central to the rational design of these materials will be a detailed understanding of the relationship between microscale network topologies and macroscale mechanics. The aim of this thesis is to characterize the relationship between the network crosslinking density and macroscale mechanics of a hydrogel system built from immunoglobulin domain 27 pentamers. Firstly a facile method is described whereby the number of crosslink sites per monomer network building block can be precisely tuned, and proteins subsequently expressed in a high-yield manner. This was then followed by the development of two assays to measure the unfolded protein fraction post-gelation and the crosslinking efficiency of each hydrogel species. The macroscale mechanics of each hydrogel species was then characterized rheologically. By correlating these microscale measurements with the macroscale mechanics the mechanism of translation between length scales is discussed. I propose that it is possible to rationally tune the mechanics of folded protein hydrogels by the use of precisely situated crosslink sites in the monomer building block. Furthermore I hypothesise that this tuneability is a result of different crosslink geometries causing changes in the network formation regime of the hydrogel. This leads to differences in network topologies and in turn hydrogel mechanics.