Chitosan-based hydrogels: Influence of crosslinking strategy on rheological properties.

The effect of two crosslink strategies on the preparation of chitosan-based covalent hydrogels was investigated employing the widely used thiol-ene reaction. This versatile "click" chemistry can be activated either photochemically or thermochemically. Initially, well-purified chitosan (CS, DA ∼4 %, Mw ∼580 kg mol-1) was separately functionalized with vinyl (CS-ene) or thiol (CS-SH) groups in aqueous media. Subsequently, two strategies were compared where thiol-ene reaction occurs respectively between: (S1) modified chitosans CS-ene and CS-SH, in a polymer - polymer strategy, and (S2) CS-ene and di(ethylene glycol) dithiol (dEG-(SH)2), in a polymer - molecule strategy. Both crosslinking strategies were evaluated through rheological measurements, starting with entangled chitosan solutions. The difference in diffusion of functional groups, whether attached to polymer chains or to free molecules, leads to faster gelation kinetics with S2. Consequently, stronger gels were obtained with S2, where the modulus was connected with the degree of functionalization, while S1 produced weaker gels closer to the percolation point, where crosslinked density was associated with the entanglement number derived from the initial concentration. Nevertheless, networks formed by both strategies were homogenous with minimal dissipative contributions to their rheological properties, indicating that structural defects are negligible.

Triblock Proteins with Weakly Dimerizing Terminal Blocks and an Intrinsically Disordered Region for Rational Design of Condensate Properties.

Condensates are molecular assemblies that are formed through liquid-liquid phase separation and play important roles in many biological processes. The rational design of condensate formation and their properties is central to applications, such as biosynthetic materials, synthetic biology, and for understanding cell biology. Protein engineering is used to make a triblock structure with varying terminal blocks of folded proteins on both sides of an intrinsically disordered mid-region. Dissociation constants are determined in the range of micromolar to millimolar for a set of proteins suitable for use as terminal blocks. Varying the weak dimerization of terminal blocks leads to an adjustable tendency for condensate formation while keeping the intrinsically disordered region constant. The dissociation constants of the terminal domains correlate directly with the tendency to undergo liquid-liquid phase separation. Differences in physical properties, such as diffusion rate are not directly correlated with the strength of dimerization but can be understood from the properties and interplay of the constituent blocks. The work demonstrates the importance of weak interactions in condensate formation and shows a principle for protein design that will help in fabricating functional condensates in a predictable and rational way.

Highly Hydrophobic Films of Engineered Silk Proteins by a Simple Deposition Method.

Molecular engineering of protein structures offers a uniquely versatile route for novel functionalities in materials. Here, we describe a method to form highly hydrophobic thin films using genetically engineered spider silk proteins. We used structurally engineered protein variants containing ADF3 and AQ12 spider silk sequences. Wetting properties were studied using static and dynamic contact angle measurements. Solution conditions and the surrounding humidity during film preparation were key parameters to obtain high hydrophobicity, as shown by contact angles in excess of 120°. Although the surface layer was highly hydrophobic, its structure was disrupted by the added water droplets. Crystal-like structures were found at the spots where water droplets had been placed. To understand the mechanism of film formation, different variants of the proteins, the topography of the films, and secondary structures of the protein components were studied. The high contact angle in the films demonstrates that the conformations that silk proteins take in the protein layer very efficiently expose their hydrophobic segments. This work reveals a highly amphiphilic nature of silk proteins and contributes to an understanding of their assembly mechanisms. It will also help in designing diverse technical uses for recombinant silk.

Intertwining Roles of the Disperse Phase Properties during Emulsification.

The combined effect of viscosity ratio, interfacial tension, and disperse phase density on the process of droplet formation during emulsification was evaluated. For that aim, emulsification by ultrasonication of oil/water systems with viscosity ratios between 1 and 600, with and without surfactant was performed. The time evolution of the average droplet size was estimated by dynamic light scattering measurements. For viscosity ratios between 1 and 200 in the presence of surfactant, our results partly reproduce those of the intriguing U-type reported in the literature. Beyond that range, the droplet size decreases, as the viscosity ratio rises. For surfactant-free systems, the size is slightly affected by the increase in viscosity. This complex scenario is analyzed in terms of both the individual and intertwined roles of interfacial tension, viscosity, and density ratios: (1) if the interfacial tension dominates, the droplet rupturing process is independent of its internal properties, and inversely, (2) if the interfacial tension is low, the internal properties play a major role in the rupturing of the droplet. Finally, we identified a scenario in which the retarded addition of surfactant leads to emulsions with a stability similar to those with the surfactant added at the beginning, saving energy and time.