Template-assembled triple-helical peptide molecules: Mimicry of collagen by molecular architecture and integrin-specific cell adhesion

Abstract

Most proteins fold into specific structures to exert their biological functions, and therefore the creation of protein-like molecular architecture is a fundamental prerequisite toward realizing a novel biologically active protein-like biomaterial. To do this with an artificial collagen, we have engineered a peptide template characterized by its collagen-like primary structure composed of Gly-Phe-Gly-Glu-Glu-Gly sequence to assemble (Pro-Hyp-Gly)n (n = 3 and 5) into triple-helical conformations that resemble the native structure of collagen. The peptide template has three carboxyl groups connected to the N-termini of three collagen peptides. The coupling was accomplished by a simple and direct branching protocol without complex strategies. A series of biophysical studies, including melting curve analyses and CD and NMR spectroscopy, demonstrated the presence of stable triple-helical conformation in the template-assembled (Pro-Hyp-Gly)3 and (Pro-Hyp-Gly)5 solution. Conversely, nontemplated peptides showed no evidence of assembly of triple-helical structure. A cell binding sequence (Gly-Phe-Hyp-Gly-Glu-Arg) derived from the collagen α1(I) chain was incorporated to mimic the integrin-specific cell adhesion of collagen. Cell adhesion and inhibition assays and immunofluorescence staining revealed a correlation of triple-helical conformation with cellular recognition of collagen mimetics in an integrin-specific way. This study offers a robust strategy for engineering native-like peptide-based biomaterials, fully composed of only amino acids, by maintaining protein conformation integrity and biological activity. © 2008 American Chemical Society.