Transferrin cycle

Vanadate transport in the erythrocyte was shown to occur via facilitated diffusion in erythrocyte membranes and was inhibited by 4,4 -diisothiocyanostilbene-2,2 -disulfonic acid (DIDS), a specific inhibitor of the band 3 anion transport protein [23], Vanadium is also believed to enter cells as the vanadyl ion, presumably through cationic facilitated diffusion systems. The divalent metal transporter 1 protein (called DMT1, and also known as Nramp2), which carries iron into cells in the gastrointestinal system and out of endosomes in the transferrin cycle [24], has been proposed to also transport the vanadyl cation. In animal systems, specific transport protein systems facilitate the transport of vanadium across membranes into the cell and between cellular compartments, whereas the transport of vanadium through fluids in the organism occurs via binding to proteins that may not be specific to vanadium. 

Kilhsch, 1. Steinlein, P. Rbmisch, K. HoUinshead, R. Beug, H. Griffiths, G. Characterization of early and late endocytic compartments of the transferrin cycle transferrin receptor antibody blocks erythroid differentiation by trapping the receptor in the early endosome. J. CeU Sci. 1992, 103,211-232. 

Figure 5.14 The transferrin to cell cycle. (From Crichton, 1991.)...

Figure 7.13 The transferrin to cell cycle HOLO-TF, diferric transferrin TFR, transferrin receptor DMT1, divalent metal transporter. (From Andrews, 2000. Reproduced by permission of Nature Reviews Genetics.)...

Once in the serum, aluminium can be transported bound to transferrin, and also to albumin and low-molecular ligands such as citrate. However, the transferrrin-aluminium complex will be able to enter cells via the transferrin-transferrin-receptor pathway (see Chapter 8). Within the acidic environment of the endosome, we assume that aluminium would be released from transferrin, but how it exits from this compartment remains unknown. Once in the cytosol of the cell, aluminium is unlikely to be readily incorporated into the iron storage protein ferritin, since this requires redox cycling between Fe2+ and Fe3+ (see Chapter 19). Studies of the subcellular distribution of aluminium in various cell lines and animal models have shown that the majority accumulates in the mitochondria, where it can interfere with calcium homeostasis. Once in the circulation, there seems little doubt that aluminium can cross the blood-brain barrier. 

A biological example of E° is the reduction of Fe(III) in the protein transferrin, which was introduced in Figure 7-4. This protein has two Fe(III)-binding sites, one in each half of the molecule designated C and N for the carboxyl and amino terminals of the peptide chain. Transferrin carries Fe(III) through the blood to cells that require iron. Membranes of these cells have a receptor that binds Fe(III)-transferrin and takes it into a compartment called an endosome into which H is pumped to lower the pH to —5.8. Iron is released from transferrin in the endosome and continues into the cell as Fe(II) attached to an intracellular metal-transport protein. The entire cycle of transferrin uptake, metal removal, and transferrin release back to the bloodstream takes 1-2 min. The time required for Fe(III) to dissociate from transferrin at pH 5.8 is —6 min, which is too long to account for release in the endosome. The reduction potential of Fe(IH)-transferrin at pH 5.8 is E° = —0.52 V, which is too low for physiologic reductants to reach. 

In the bloodstream, ferric iron binds tightly to circulating plasma transferrin (TF) to form diferric transferrin (FeTF). Absorption of iron into erythrocytes depends on basolateral membrane receptor-mediated endocytosis of FeTF by transferrin receptor 1 (TfR 1). FeTF binds to TfR 1 on the surface of erythroid precursors. These complexes invaginate in pits on the cell surface to form endosomes. Proton pumps within the endosomes lower pH to promote the release of iron into the cytoplasm from transferrin. Once the cycle is completed,TF and TfR 1 are recycled back to the cell surface. TF and TfR 1 play similar roles in iron absorption at the basolateral membrane of crypt enterocytes (Parkilla et al., 2001 Pietrangelo, 2002). 

The dwell time of a transferrin molecule with the reticulocyte may be only a minute or two (52, 80) or possibly as long as 10 min (86), after which the protein is released for another cycle of iron transport. The affinity of reticulocyte receptors for apotransferrin appears less than for iron-loaded molecules (80). This property may facilitate the release of the protein from the receptor after it has donated its iron and ensures that the apoprotein will not impede the delivery of iron to reticulocytes by competing with iron-bearing molecules for available receptors on the cell surface. 

Fig. 1. Schematic representation of the cycle of iron delivery to cells by transferrin, showing the uptake of diferric transferrin by cell receptors, internalization, release of iron at the lower intracellular pH, and recycling and release of apotransferrin.

The most striking feature of transferrin chemistry is that iron is bound with extraordinary avidity, yet it can be released without any denaturation and the protein can be recycled through many cycles of uptake and release. The mechanisms by which this is done are of fundamental importance to understanding biological transport processes. 

Figure 4. Cycle-1 chromatograms of polyproline (10 nmols), ovalbumin (5 nmols), and apo-transferrin (5 nmols) showing the identification of the C-terminal thiohydantoin-proline residue. Approximate 10-20% initial recoveries are found.

Carbonic anhydrase and transferrin immobilized onto membranes either by ProSorb or by SDS-PAGE/electroblottig were subjected to 10-20 cycles of Edman degradation on an Applied Biosystems 473 Sequencer. The sequenced membranes were used direcdy for successive chemical fragmentation. 

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