Strong, Tough, Stretchable, and Self-Adhesive Hydrogels from Intrinsically Unstructured Proteins.

Strong, tough, stretchable, and self-adhesive hydrogels are designed with intrinsically unstructured proteins. The extraordinary mechanical properties exhibited by these materials are enabled by an integration of toughening mechanisms that maintain high elasticity and dissipate mechanical energy within the protein networks.

Simple assay for proteases based on aggregation of stimulus-responsive polypeptides.

Unregulated changes in protease activity are linked to many diseases including cancer. Fast, accurate, and low-cost assays for detection of these changes are being explored for early diagnosis and monitoring of these diseases and can also be used as platforms for the discovery of new drugs. We report a new methodology for the simple detection and quantification of protease activity in buffer and human serum. The assay is based on recombinant diblock polypeptides that undergo temperature- or salt-triggered micellization in water. The coronae of the micelles are linked to the water-insoluble cores by a peptide substrate that is cleaved in the presence of the target protease. Protease cleavage of the diblock polypeptide triggers the aggregation of the core-forming segment, leading to a change in solution optical density, which can be used to detect the presence of, and to quantify the concentration of, protease. We used matrix metalloproteinase-1 (MMP-1) as a model protease and found peptide aggregation time to be proportional to enzyme concentration over a range from endogenous MMP-1 level in human serum (∼3 ng/mL) to 100 ng/mL (0.15-5 nM) in 40% human serum and 1-100 ng/mL in buffer. The assay does not require any intermediate steps or sophisticated data analysis, and the modular design of the assay system is amenable to straightforward adaptation for the detection of a wide range of proteases.

Size and shape characterization of thermoreversible micelles of three-armed star elastin-like polypeptides.

Three-armed star elastin-like polypeptides are shown to have the capability of self-assembling into micellar constructs at certain environmental conditions. Here, a study of the size distribution, shape, and molecular weight of these micelles at different salt concentrations and pH values is presented. Multiangle dynamic light scattering was used to study the formation, reversibility, and size of the micelles at different environmental conditions. On the basis of the salt concentration of the solution, two distinct size distribution regimes and a transition region were observed. Static light scattering was performed to study the molecular weight and geometrical anisotropy of the micelles in each regime. The anisotropic behavior and elongation of the particles were independently confirmed by depolarized dynamic light scattering, and a model for micelles at each regime was proposed. The size and molecular weight of the micelles were verified using viscosity measurements. The results of this study suggest that there is big jump in the size and molecular weight of the micelles from the first salt-dependent regime to the other, and the shape of the micelles changes from spheres to cylindrical micelles with a higher than 10:1 axis ratio.

Molecular architecture influences the thermally induced aggregation behavior of elastin-like polypeptides.

Elastin-like polypeptides are thermally responsive polymers that exhibit phase separation above a transition temperature. The effect of molecular architecture on the temperature responsive behavior of elastin-like polypeptide solutions was investigated by characterization of solutions of three-armed star polypeptides, linear polypeptides, and their mixtures. These biosynthesized polypeptides have precise lengths and amino acid sequences. Transition temperatures were measured as a function of molecular weight and solution concentration and compared to their linear counterparts. Like their linear counterparts, the transition temperature is linearly related to log concentration. A mathematical relationship was used to fit the transition temperature data of different polypeptide lengths to a volume-based concentration using the polymer coil volume. The results of this model suggest that the linear ELP is in a random coil conformation at the transition temperature while the three-armed ELP is in a compact extended coil conformation, consistent with different pathways for aggregation. Solutions containing both trimer and linear constructs have two transition temperatures, further supporting differing aggregation behaviors.