The very high binding affinity of
avidin to
biotin is one of the highest to occur in nature. We constructed a fusion
protein composed of
avidin and the endocytotic
LDL receptor in order to target biotinylated molecules to cells of the desired tissues. In addition to the native
avidin, charge-mutated and nonglycosylated
avidins were utilized as part of the fusion
proteins, in order to modify its properties. All of the fusion
protein versions retained the
biotin-binding capacity. Although the specificity was not increased, however, fusion
proteins composed of natural
avidin and nonglycosylated
avidin bound most efficiently to the biotinylated
ligands. Fluorescence microscopy and atomic force microscopy studies revealed the expression of the fusion
protein on cell membranes, and demonstrated specific and high-affinity binding of
biotin to the
low-density lipoprotein receptor (LDLR)-
avidin fusion
protein in vitro. Additionally, systemically administered biotinylated
ligand targeted with high specificity the intracerebral
tumors of rats that were expressing fusion
protein after the virus-mediated gene transfer. These results suggest that local gene transfer of the fusion
protein to target tissues may offer a novel tool for the delivery of biotinylated molecules in vitro and in vivo for therapeutic and imaging purposes.