As substrates for cell encapsulation, protein hydrogels have potential applications in tissue regeneration, wound healing, and stem cell differentiation. Protein networks are also viable platforms for studying cell growth, cell migration, and cell interactions in an environment that mimics the extracellular matrix.  Molecular network defects, such as dangling chain ends and elastically inactive primary loops, may impact the behavior of cells in currently unknown ways; correlation between these defects and cell migration would provide new molecular insights into cell/biomaterial interactions, and define the shortest critical length scales for hydrogel design.  Herein, we show that the fraction of primary loops in an artificial protein network can be quantified using a technique called network disassembly spectrometry (NDS).  In this approach, an artificial random coil protein macromer is recombinantly expressed and chemically crosslinked to form hydrogels. Amino acid sequences at key positions within the macromer encode enzymatically and chemically degradable domains.  Degradation of the material followed by HPLC and UV-Vis spectroscopic analysis of the network disassembly products allows for determination of the fraction of primary loops and dangling chains in the protein hydrogel (Figure 1).  Comparison of NDS data for cell-induced degradation versus chemical degradation provides direct insight into interactions between cells and molecular network defects.     

Measuring primary loop fractions in protein hydrogels

247th National Meeting (2014)

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