The Transferrins

A variety of cellular and viral proteins contain fatty acids covalently bound via ester linkages to the side chains of cysteine and sometimes to serine or threonine residues within a polypeptide chain (Figure 9.18). This type of fatty acyl chain linkage has a broader fatty acid specificity than A myristoylation. Myristate, palmitate, stearate, and oleate can all be esterified in this way, with the Cjg and Cjg chain lengths being most commonly found. Proteins anchored to membranes via fatty acyl thioesters include G-protein-coupled receptors, the surface glycoproteins of several viruses, and the transferrin receptor protein. 

Synthesis of the transferrin receptor (TfR) and that of ferritin are reciprocally linked to cellular iron content. Specific untranslated sequences of the mRNAs for both proteins (named iron response elements) interact with a cytosolic protein sensitive to variations in levels of cellular iron (iron-responsive element-binding protein). When iron levels are high, cells use stored ferritin mRNA to synthesize ferritin, and the TfR mRNA is degraded. In contrast, when iron levels are low, the TfR mRNA is stabilized and increased synthesis of receptors occurs, while ferritin mRNA is apparently stored in an inactive form. This is an important example of control of expression of proteins at the translational level. 

HFE has been shown to be located in cells in the crypts of the small intestine, the site of iron absorption. There is evidence that it associates with P2 niicroglobu-lin, an association that may be necessary for its stability, intracellular processing, and cell surface expression. The complex interacts with the transferrin receptor (TfR) how this leads to excessive storage of iron when HFE is altered by mutation is under close smdy. The mouse homolog of HFE has been knocked out, resulting in a potentially useful animal model of hemochromatosis. 

Figure 5.10 Ribbon diagram of the transferrin receptor dimer depicted in its likely orientation with regard to the plasma membrane. One monomer is blue, the other is coloured according to domain the protease-like, apical and helical domains are red, green and yellow respectively the stalk is shown in grey, connected to the putative membrane spanning helices in black. Pink spheres indicate the location of Sm3+ ions. Reprinted with permission from Lawrence et ah, 1999. Copyright (1999) American Association for the Advancement of Science.

We will deal here only with proteins of class (ii) that contain diiron centres and reserve the discussion of proteins containing polyiron oxo aggregates (ferritins) until Chapter 6. As for the proteins of class (iii), their discussion is deferred until Chapter 3 concerning siderophore receptors and Chapter 5 concerning the transferrins and their receptors respectively. 

The determination of the structure of the iron transporter, ferric-binding, protein (hFBP)t from Haemophilus influenzae (Bruns et ah, 1997) at 0.16 nm resolution shows that it is a member of the transferrin superfamily, which includes both the transferrins and a number of periplasmic binding proteins (PBP). The PBPs transport a wide variety of nutrients, including sugars, amino acids and ions, across the periplasm from the outer to the inner (plasma) membrane in bacteria (see Chapter 3). Iron binding by transferrins (see below) requires concomitant binding of a carbonate anion, which is located at the N-terminus of a helix. This corresponds to the site at which the anions are specifically bound in the bacterial periplasmic sulfate- and... 

Figure 5.3 The deduced evolutionary tree for selected members of the transferrin superfamily, based on comparisons of structures and sequences. The tree combines the transferrins with a number of prokaryotic periplasmic transport proteins. From Bruns et al., 1997. Reproduced by permission of Nature Publishing Group.

With the advent of monoclonal antibodies, the search for tumour-specific antigens became the biggest cottage industry since unemployment. It rapidly became apparent that a 90 kD disulfide-bridged transmembrane protein was present in many tumour cells - it was the transferrin receptor, and as they say, the rest is history. It has become a standard procedure to determine the in vivo growth potential of tumours by measuring transferrin receptor expression. 

Figure 5.9 Diagrammatic representation of the transferrin receptor. Adapted from Aisen, 1998, by courtesy of Marcel Dekker, Inc.

The equivalent of the tryptic fragment of human transferrin receptor has been expressed in Chinese hamster ovary cells and its structure determined at a resolution of 0.32 nm (Lawrence et ah, 1999). The asymmetric unit of the crystals contains four transferrin receptor dimers. Interpretable electron density is found for the entire tryptic fragment except for Arg-121 at the amino terminus, and density is also seen for the first N-acetylglucosamine residue at each of the N-glycosylation sites. The transferrin receptor monomer is made up of three distinct domains, organized such that the dimer is butterfly shaped (Figure 5.10, Plate 7). The likely orientation of the dimer with respect to the plasma membrane has been assigned on the basis of the... 

A human cDNA highly homologous to the transferrin receptor, has been identified and reported to encode a protein, designated TfR2, which binds diferrictransferrin and mediates iron uptake of transferrin-bound iron (Kawabata etal., 1999). A mouse orthologue of human TfR2 has been found independently (Fleming etal.,... 

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

Another potential source of iron, at least for hepatocytes, is receptor-independent uptake of iron from transferrin. This appears to involve an iron uptake pathway from transferrin which is neither suppressed in hepatocytes by antibodies to TfR (Trinder et at, 1988), nor by transfection of HuH-7 hepatoma cells with transferrin receptor anti-sense cDNA (Trinder etat, 1996). The same pathway may also be utilized for iron uptake from isolated transferrin N-lobe, which is not recognized by the receptor (Thorstensen et at, 1995). The possible role of TfR2 in this process remains to be established, as does the physiological importance of this pathway in intact liver. Human melanoma cells (Richardson and Baker, 1994) and Chinese hamster cells lacking transferrin receptors but transfected with melanotransferrin (Kennard et at, 1995) use another pathway for transferrin iron uptake, independent of the transferrin receptor, but utilizing iron transfer from transferrin or simple iron chelates to membrane-anchored melanotransferrin, and from there onwards into the cellular interior. 

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