Helen Waller
Newcastle University, UK
Title: Engineered Caf1 protein polymers form tuneable bioactive hydrogel scaffolds
Biography
Biography: Helen Waller
Abstract
Here, we describe a new animal-free biomaterial with potential uses in 3D-tissue culture and regenerative medicine due to its low cost, high stability and definable bioactivity. Capsular antigen fraction1 (Caf1) is a protein from the plague bacterium Yersinia pestis that is secreted via the chaperone-usher pathway and protects the pathogen from phagocytosis by forming a non-stick protective layer around the cell. The 15.5 kDa monomer has an Ig-like fold and resembles the extracellular matrix protein fibronectin. The subunits polymerize via donor-strand complementation, forming a highly stable non-covalent polymer. In this work, recombinant Caf1 polymers produced via batch fermentation using Escherichia coli were secreted by the bacterium into a flocculent layer above the cell pellet, and could be easily extracted and purified in large quantities. We demonstrate the polymers robust thermostability by circular dichroism and SDS-PAGE, and observe their large size using electron microscopy and SEC-MALS. Additionally, we have selectively reversed the natural non-stick behaviour of the WT polymer by introducing an integrin binding sequence, RGDS, into loop 5 that can promote U2OS cell adhesion. Additional bioactive peptides motifs from Osteopontin, bone morphogenic protein 2, collagen and laminin were then introduced at different positions within Caf1. Finally, PEG based chemical cross linkers were used to form stable 3D hydrogels with designed porosities and tuneable stiffness, ideal for use in cell culture and drug delivery applications. The combination of these new motifs into tuneable Caf1 hydrogels will help to expand the functionality of this exciting new biomaterial for use in a variety of biomedical applications.