Dynamic Presentation of Immobilized Ligands Regulated through Biomolecular Recognition

To mimic the dynamic regulation of signaling ligands immobilized on extracellular matrices or on the surfaces of neighboring cells for guidance of cell behavior and fate selection, we have harnessed biomolecular recognition in combination with polymer engineering to create dynamic surfaces on which the accessibility of immobilized ligands to cell surface receptors can be reversibly interconverted under physiological conditions. The cell-adhesive RGD peptide is chosen as a model ligand. RGD is fused to the C-terminus of a leucine zipper domain A, and this fusion polypeptide is immobilized on surfaces through a residue at the N-terminus. The immobilized RGD can be converted from a cell-accessible to a cell-inaccessible state by addition of a conjugate of poly(ethylene) glycol (PEG) and another leucine zipper domain B (B-PEG). Heterodimerization between A and B allows coimmobilization of the PEG, which shields RGD from access by cells. The shielded RGD can be converted back to a cell-accessible state by addition of nonimmobilized polypeptide A, which competes with the immobilized A for binding to B-PEG and removes B-PEG from the surface. This molecular design offers several advantages: the interconversion is reversible; the ligand remains immobilized during dynamic regulation so that cells are not exposed to the soluble form of the ligand that potentially has detrimental effects; the precision of the on/off states is assured by the molecular-level uniformity of the ligand and PEG coimmobilized through leucine zipper heterodimerization. The method can be readily adapted for dynamic regulation of other immobilized bioactive ligands of interest.