The
coiled-coil folding motif represents an ideal scaffold for the defined
presentation of ligands due to the possibility of positioning them
at specific distances along the axis. We created a coiled-coil glycopeptide
library to characterize the distances between the carbohydrate-binding
sites of the asialoglycoprotein receptors (ASGPR) on hepatocytes.
The components of the glycopeptide library vary for the number of
displayed ligands (galactose), their position on the peptide sequence,
and the space between peptide backbone and carbohydrate. We determined
the binding of the glycopeptides to the hepatocytes, and we established
the optimal distance and orientation of the galactose moieties for
interaction with the ASGPR using flow cytometry. We confirmed that
the binding occurs through endocytosis mediated by ASGPR <i>via</i> inhibition studies with cytochalasin D; fluorescence microscopy
studies display the uptake of the carrier peptides inside the cell.
Thus, this study demonstrates that the coiled-coil motif can be used
as reliable scaffold for the rational presentation of ligands