(448a) Elucidating Human Transferrin-Receptor Interactions Using Linear Epitope Mapping

Transferrin (Tf) is a carrier protein involved in the transport of iron (Fe3+). Tf binds to transmembrane transferrin receptors (TfRs) expressed on cell surfaces and transports Fe3+ by receptor mediated transcytosis (RMT). The Tf-TfR complex is an interesting system as it provides a natural biological pathway for delivery of Tf-bound cargo across cells and tissues. It has also been demonstrated that cancer cells over-expressing TfR can be selectively targeted with Tf-conjugated cytotoxic drugs. Understanding the interactions between Tf and TfR is therefore valuable for designing better therapeutic modalities for a broad range of physiological diseases. A few studies have highlighted putative amino acid residues involved in the formation of the Tf-TfR complex. The crystal structure of the N-lobe of Tf and the ectodomain of TfR has also been resolved independently. However, the crystal structure of the bound human Tf-TfR complex is still not available. We propose to use peptide microarrays for epitope mapping of the Tf-TfR complex to probe the interactions between Tf and TfR. Linear epitope mapping has been widely used as a technique for identifying antigenic regions in proteins in a fast, reliable high-throughput fashion. We have employed peptide microarrays of short peptides that are linear, continuous fragments of TfR to capture the interactions between TfR and its ligand. Microarrays printed with peptide fragments that span the entire sequence of TfR were tested against Tf to screen for regions on TfR that are involved in binding to Tf. Our results have confirmed prior findings but also highlighted the role of other important discontinuous regions on Tf involved in binding to TfR. We anticipate that our studies along with existing knowledge about the Tf-TfR system will further advance the understanding about this important transport and regulatory pathway.