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Exploring Transferrin-Receptor Interactions at the Single-Molecule Level

Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, Yokohama, Kanagawa, Japan

Correspondence: Address reprint requests to Alexandre Yersin, Tokyo Institute of Technology, Graduate School of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259-B8 Nagatsuta-cho, Midori-ku, Yokohama 226-8501, Japan. Tel.: 81-45-924 5129; E-mail: ayersin{at}bio.titech.ac.jp.

Interaction between the iron transporter protein transferrin (Tf) and its receptor at the cell surface is fundamental for most living organisms. Tf receptor (TfR) binds iron-loaded Tf (holo-Tf) and transports it to endosomes, where acidic pH favors iron release. Iron-free Tf (apo-Tf) is then brought back to the cell surface and dissociates from TfR. Here we investigated the Tf-TfR interaction at the single-molecule level under different conditions encountered during the Tf cycle. An atomic force microscope tip functionalized with holo-Tf or apo-Tf was used to probe TfR. We tested both purified TfR anchored to a mica substrate and in situ TfR at the surface of living cells. Dynamic force measurements showed similar results for TfR on mica or at the cell surface but revealed striking differences between holo-Tf-TfR and apo-Tf-TfR interactions. First, the forces necessary to unbind holo-Tf and TfR are always stronger compared to the apo-Tf–TfR interaction. Second, dissociation of holo-Tf-TfR complex involves overcoming two energy barriers, whereas the apo-Tf-TfR unbinding pathway comprises only one energy barrier. These results agree with a model that proposes differences in the contact points between holo-Tf-TfR and apo-Tf-TfR interactions.