In spite of these large conformational changes, apohTF remains bound to the TFR at the mildly acidic pH (~5.6) of the endosome, implying that the binding partners accommodate and compensate for these structural changes ( 7). Large conformational changes in each lobe are associated with opening of the cleft and iron release ( 6). Within the endosome, following clathrin-dependent endocytosis of the hTF/TFR complex and upon exposure to a slightly more acidic pH, hTF releases Fe 3+ to an unidentified chelator in a TFR mediated process. A transmembrane glycoprotein, each TFR monomer is composed of three domains: a helical domain responsible for dimerization, as well as an apical and protease-like domain ( 4).Īt pH 7.4, the TFR preferentially binds diferric hTF (with low nM affinity), the two monoferric hTFs bind ~10 fold weaker, and apohTF binds very weakly, if at all ( 5). Physiologically, one molecule of hTF binds tightly to each monomer of the homodimeric transferrin receptor (TFR) located on the extracellular surface of dividing cells. Four unequally distributed species of hTF differing with regard to iron content are found in plasma: diferric hTF, monoferric N-lobe hTF, monoferric C-lobe hTF, and apohTF (iron-free) ( 1- 3). Sequestration of highly insoluble Fe 3+ by hTF maintains iron in the blood in a non-reactive state, preventing reduction to ferrous iron (Fe 2+) which can catalyze the production of reactive oxygen species via Fenton chemistry. The homologous N- and C-lobes of hTF are divided into two subdomains (N1 and N2, C1 and C2), that fold to form a deep cleft capable of binding a single ferric iron. The transport of iron throughout the body by human serum transferrin (hTF) 1 is central to iron homeostasis. Thus, mutagenesis of charged hTF residues has allowed identification of a number of residues that are critical to formation of and iron release from the hTF/TFR complex.
![Htf Digiter 1.0 Htf Digiter 1.0](https://a.deviantart.net/avatars-big/d/i/digiguardia.jpg)
Moreover, mutation of three residues (Asp356, Glu367 and Lys511), whether in the diferric or monoferric C-lobe hTF, significantly affected iron release when in complex with the TFR. In particular, we show that Asp356 in the C-lobe of hTF is essential to the formation of a stable hTF/TFR complex: mutation of Asp356 in the monoferric C-lobe hTF background prevented the formation of the stoichiometric 2:2 (hTF:TFR monomer) complex.
![Htf Digiter 1.0 Htf Digiter 1.0](https://t00.deviantart.net/vT9oXJ3pqnwvf1wrCAF81ltygio=/fit-in/700x350/filters:fixed_height(100,100):origin()/pre00/5d33/th/pre/i/2010/025/a/4/fang_by_flaky013.jpg)
Six hTF mutants (R50A, R352A, D356A, E357A, E367A and K511A) competed poorly with biotinylated diferric hTF for binding to TFR. Alanine substitution of eleven charged hTF residues identified by available structures and modeling studies allowed evaluation of the role of each in (1) binding of hTF to the TFR and (2) in TFR-mediated iron release. Identification of the specific residues accounting for the pH-sensitive nanomolar affinity with which hTF binds to TFR throughout the cycle is important to fully understand the iron delivery process. The return of hTF to the blood to continue the iron delivery cycle relies on the maintenance of the interaction between apohTF and the TFR after exposure to endosomal pH (≤ 6.0). Internalization of the complex into an endosome precedes iron removal. Efficient delivery of iron is critically dependent on the binding of diferric human serum transferrin (hTF) to its specific receptor (TFR) on the surface of actively dividing cells.