Acetic Acid Enables Precise Tailoring of the Mechanical Behavior of Protein-Based Hydrogels.

Engineering viscoelastic and biocompatible materials with tailored mechanical and microstructure properties capable of mimicking the biological stiffness (<17 kPa) or serving as bioimplants will bring protein-based hydrogels to the forefront in the biomaterials field. Here, we introduce a method that uses different concentrations of acetic acid (AA) to control the covalent tyrosine-tyrosine cross-linking interactions at the nanoscale level during protein-based hydrogel synthesis and manipulates their mechanical and microstructure properties without affecting protein concentration and (un)folding nanomechanics. We demonstrated this approach by adding AA as a precursor to the preparation buffer of a photoactivated protein-based hydrogel mixture. This strategy allowed us to synthesize hydrogels made from bovine serum albumin (BSA) and eight repeats protein L structure, with a fine-tailored wide range of stiffness (2-35 kPa). Together with protein engineering technologies, this method will open new routes in developing and investigating tunable protein-based hydrogels and extend their application toward new horizons.

Kinetic Method of Producing Pores Inside Protein-Based Biomaterials without Compromising Their Structural Integrity.

Hydrogels made from globular proteins cross-linked covalently into a stable network are becoming an important type of biomaterial, with applications in artificial tissue design and cell culture scaffolds, and represent a promising system to study the mechanical and biochemical unfolding of proteins in crowded environments. Due to the small size of the globular protein domains, typically 2-5 nm, the primary network allows for a limited transfer of protein molecules and prevents the passing of particles and aggregates with dimensions over 100 nm. Here, we demonstrate a method to produce protein materials with micrometer-sized pores and increased permeability. Our approach relies on forming two competing networks: a covalent network made from cross-linked bovine serum albumin (BSA) proteins via a light-activated reaction and a physical network triggered by the aggregation of a polysaccharide, alginate, in the presence of Ca2+ ions. By fine-tuning the reaction times, we produce porous-protein hydrogels that retain the mechanical characteristics of their less-porous counterparts. We further describe a simple model to investigate the kinetic balance between the nucleation of alginate and cross-linking of BSA molecules and find the upper rate of the alginate aggregation reaction driving pore formation. By enabling a more significant permeability for protein-based materials without compromising their mechanical response, our method opens new vistas into studying protein-protein interactions and cell growth and designing novel affinity methods.

Cation-induced shape programming and morphing in protein-based hydrogels.

Smart materials that are capable of memorizing a temporary shape, and morph in response to a stimulus, have the potential to revolutionize medicine and robotics. Here, we introduce an innovative method to program protein hydrogels and to induce shape changes in aqueous solutions at room temperature. We demonstrate our approach using hydrogels made from serum albumin, the most abundant protein in the blood plasma, which are synthesized in a cylindrical or flower shape. These gels are then programmed into a spring or a ring shape, respectively. The programming is performed through a marked change in stiffness (of up to 17-fold), induced by adsorption of Zn2+ or Cu2+ cations. We show that these programmed biomaterials can then morph back into their original shape, as the cations diffuse outside the hydrogel material. The approach demonstrated here represents an innovative strategy to program protein-based hydrogels to behave as actuators.