Self-assembling protein hydrogels with modular integrin binding domains.
Hydrogels with integrin binding activity were created from associating proteins with embedded RGD sequences. These proteins are a modified AC(10)Bcys 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 AC(10)Bcys 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 AC(10)Bcys 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.
Mi
Lixin
L
Department of Chemical & Biomolecular Engineering, Johns Hopkins University, Baltimore, MD 21218, USA.
Fischer
Stephen
S
Chung
Brian
B
Sundelacruz
Sarah
S
Harden
James L
JL
D000595:Amino Acid Sequence; D002448:Cell Adhesion; D002478:Cells, Cultured; D002942:Circular Dichroism; D004548:Elasticity; D005347:Fibroblasts; D006801:Humans; D020100:Hydrogels / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D016023:Integrins / Q000378:metabolism*; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry* / Q000378:metabolism*; D014783:Viscosity
D020100:Hydrogels; D016023:Integrins; D009842:Oligopeptides; D011506:Proteins; C047981:arginyl-glycyl-aspartic acid