Protein Engineered Triblock Polymers Composed of Two SADs: Enhanced Mechanical Properties and Binding Abilities.

Recombinant methods have been used to engineer artificial protein triblock polymers composed of two different self-assembling domains (SADs) bearing one elastin (E) flanked by two cartilage oligomeric matrix protein coiled-coil (C) domains to generate CEC. To understand how the two C domains improve small molecule recognition and the mechanical integrity of CEC, we have constructed CL44AECL44A, which bears an impaired CL44A domain that is unstructured as a negative control. The CEC triblock polymer demonstrates increased small molecule binding and ideal elastic behavior for hydrogel formation. The negative control CL44AECL44A does not exhibit binding to small molecule and is inelastic at lower temperatures, affirming the favorable role of C domain and its helical conformation. While both CEC and CL44AECL44A assemble into micelles, CEC is more densely packed with C domains on the surface enabling the development of networks leading to hydrogel formation. Such protein engineered triblock copolymers capable of forming robust hydrogels hold tremendous promise for biomedical applications in drug delivery and tissue engineering.

Circular Dichroistic Impacts of 1-(3-Dimethylaminopropyl)-3-ethylurea: Secondary Structure Artifacts Arising from Bioconjugation Using 1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide.

1-Ethyl-3-[3-dimethylaminopropyl]carbodiimide (EDC) is a commonly used reagent for bioconjugation and peptide synthesis. Both EDC and the corresponding urea derivative, 1-(3-dimethylaminopropyl)-3-ethylurea (EDU), are achiral. As the reagent is active in aqueous solutions, it is a common choice for the study of evolving secondary structural changes via circular dichroism. This work highlights the effect of EDU on spectropolarimetric measurements, namely, the problematic absorption profile at low wavelengths (190-220 nm). We demonstrate that EDU is capable of erroneously indicating structural changes, particularly loss of α-helical character, through masking of the characteristic minimum at 208 nm. However, if the concentrations of the EDU in the sample are known, then this effect can be anticipated and calculations of secondary structure can be adjusted to avoid the impacted wavelengths. Impacts of EDU in a sample are compared to those of standard urea, which, by contrast, is commonly used as a denaturant in circular dichroism studies without issue.

Channel density and porosity of degradable bridging scaffolds on axon growth after spinal injury.

Bridges implanted into the injured spinal cord function to stabilize the injury, while also supporting and directing axon growth. The architecture of the bridge is critical to its function, with pores to support cell infiltration that integrates the implant with the host and channels to direct axon elongation. Here, we developed a sucrose fiber template to create poly(lactide-co-glycolide) multiple channel bridges for implantation into a lateral hemisection that had a 3-fold increase in channel number relative to previous bridges and an overall porosity ranging from approximately 70%-90%. Following implantation into rat and mouse models, axons were observed within channels for all conditions. The axon density within the bridge increased nearly 7-fold relative to previous bridges with fewer channels. Furthermore, increasing the bridge porosity substantially increased the number of axons, which correlated with the extent of cell infiltration throughout the bridge. Analysis of these cell types identified an increased presence of mature oligodendrocytes within the bridge at higher porosities. These results demonstrate that channels and bridge porosity influence the re-growth of axons through the injury. These bridges provide a platform technology capable of being combined with the delivery of regenerative factors for the ultimate goal of achieving functional recovery.