Self-Assembling Protein Hydrogels with Modular Integrin Binding Domains

• Published in: Biomacromolecules 2006-01, Vol.7 (1), p.38-47
• Main Authors: Mi, Lixin, Fischer, Stephen, Chung, Brian, Sundelacruz, Sarah, Harden, James L
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Applied sciences; Biological and medical sciences; Cell Adhesion; Cells, Cultured; Circular Dichroism; Elasticity; Exact sciences and technology; Fibroblasts; Fundamental and applied biological sciences. Psychology; Humans; Hydrogels - chemistry; Hydrogen-Ion Concentration; Integrins - metabolism; Miscellaneous; Molecular biophysics; Molecular Sequence Data; Natural polymers; Oligopeptides - chemistry; Oligopeptides - metabolism; Physico-chemical properties of biomolecules; Physicochemistry of polymers; Protein Binding; Protein Structure, Secondary; Protein Structure, Tertiary; Proteins; Proteins - chemistry; Proteins - metabolism; Viscosity
• ISSN: 1525-7797; 1526-4602
• DOI: 10.1021/bm050157p

Hydrogels with integrin binding activity were created from associating proteins with embedded RGD sequences. These proteins are a modified AC10Bcys triblock design composed of acidic A and basic B leucine zipper associating domains flanking a new soluble disordered coil block that contains nine repeats of AGAGAGPEG and three copies of the RGD integrin binding sequence. As with the original AC10Bcys design without the embedded RGD sequences, these proteins self-assemble into stable hydrogels at concentrations above approximately 50 mg/mL in a range of solution pH and temperature conditions. The mechanism for hydrogel assembly is the intermolecular association of A and B helical domains into bundles which act as cross-links connected by the soluble central disordered coil domains. The secondary structure of the proteins and the mechanical properties of the hydrogels they form are not adversely affected by the presence of the RGD sequences. The RGD sequences embedded in the disordered coil region support the adhesion, spreading, and polarization of human fibroblast cells on protein coated surfaces. Confocal microscopy studies demonstrated the presence of focal adhesion complexes and organized actin stress fibers in these cells. In contrast, fibroblasts seeded onto surfaces coated with the original AC10Bcys protein remained rounded and did not form focal adhesions, indicating that bioactivity is conferred by the presence of the embedded RGD sequences. Such hydrogel-forming bioactive proteins have potential for cell and tissue culture applications.