Lactoferrin pattern

Figure 5.2 Schematic representation of the folding pattern for the N-lobe (left) and C-lobe (right) of human lactoferrin. From Anderson et al., 1989. Reproduced by permission of Academic Press.

Detailed pictures of the iron-binding sites in transferrins have been provided by the crystal structures of lactoferrin (Anderson et ai, 1987, 1989 Baker etai, 1987) and serum transferrin (Bailey etal., 1988). Each structure is organized into two lobes of similar structure (the amino- and carboxy-terminal lobes) that exhibit internal sequence homology. Each lobe, in turn, is organized into two domains separated by a cleft (Fig. 3 and 10). The domains have similar folding patterns of the a//3 type. One iron site is present in each lobe, which occupies equivalent positions in the interdomain cleft. The same sets of residues serve as iron ligands to the two sites two tyrosines, one histidine, and one aspartate. Additional extra density completes the octahedral coordination of the iron and presumably corresponds to an anion and/or bound water. The iron sites are buried about 10 A below the protein surface and are inaccessible to solvent. 

Distribution of Cu in milk has been studied by SEC coupled to ICP-AES [12, 14], and ICP-MS [15, 17-19], or even using electrothermal atomization atomic emission spectrometry (ET-AAS) for the detection [20]. Breast milk Cu seems to be distributed all over the biocompounds from high (caseins, immunoglobulins, lactoferrin, and serum albumin) to lower-molecular-weight ligands (lac-toalbumin, peptones, free aminoacids, citrates, etc.). The distribution patterns of Cu have been shown to be very different in mature milk and colostrum [18] (Fig. 17.3). 

We subsequently used extrinsically labeled diets for further fractionation by gel filtration and immunoaffinity chromatography. Mn in human milk was found to be predominantly (about 70%) bound to lactoferrin, the major iron-binding protein in human milk. Minor amounts were bound to casein in human milk and the milk fat globule membrane. It is therefore possible that changes in lactoferrin concentration in human milk may explain the developmental pattern... 

A final point of general organization concerns the carbohydrate. All transferrins so far characterized, except apparently for one fish transferrin (84), are glycoproteins. There is, however, no pattern to the sites of attachment of the carbohydrate chains on different proteins—they appear almost randomly distributed over the protein surface (Fig. 4), strengthening the view that the carbohydrate plays no direct role in function. Rabbit serum transferrin, for example, has one carbohydrate chain, on its C-lobe (residue 490) human serum transferrin has two, both on the C-lobe (residues 416 and 611) human lactoferrin has two, one on each lobe (at residues 137 and 478) and bovine lactoferrin has four, one on the N-lobe (residue 233) and three on the C-lobe (residues 368, 476, and 545). 

Fig. 5. Polypeptide folding pattern found in each lobe of human lactoferrin. Helices (cylinders) are numbered 1 to 12 and /3-strands (arrows) are labeled a to k as in Anderson et al. (78). The interdomain backbone strands are shaded and the position of the hinge is indicated.

Most remarkably, one group of the bacterial binding proteins, which includes the two anion-binding proteins so far analyzed [specific for sulfate (115) and phosphate (116)], has even closer similarity. First, the polypeptide folding pattern in these proteins is almost identical to that in each lobe of lactoferrin (Fig. 15) the central /3-sheet of each... 

Fig. 15. Polypeptide folding patterns for (a) one-half of a transferrin molecule (the N-lobe of lactoferrin) and (b) the bacterial periplasmic sulfate-binding protein. Adapted from Baker et al. (85), with permission.

The recombinant whole molecules are both expressed in glycosylated form, although the glycosylation patterns differ from the proteins isolated from natural sources. The recombinant human transferrin binds to receptors both in its glycosylated form and as a nonglycosylated mutant, showing that the carbohydrate is not required for receptor binding (230). Recombinant human lactoferrin shows identical spectroscopic properties and shows an identical profile of pH-dependent iron release when compared with human milk lactoferrin (231). 

Three-dimensional structure of lactotransferrin. Top schematic representation of the folding pattern of each lactoferrin lobe Domain I is based on a beta-sheet of four parallel and two antiparallel domains Domain II is formdd from four parallel and one antiparallel strand. Bottom stereo Ca diagram of the N lobe of lactoferrin ( ) iron atom between domain I (residues 6-90-I-) and domain II (residues 91-251) ( ) disulfide bridges ( ) carbohydrate attachment site. See Reference 39. 

Two-dimensional electrophoretic analysis of tear proteins followed by immunoblotting has shown that the major components are lactoferrin, lysozyme, albumin, secretory IgA, and five tear-specific proteins. There are significant differences in the amounts of lactoferrin and two kinds of tear-specific proteins differ between the sexes. A comparison of the two-dimensional electrophoretic patterns of tear proteins from patients with conjunctivitis with normal individuals shows reductions in the levels of IgA, lactoferrin, and tear-specific proteins whilst the intensity of albumin is increased in addition, haptoglobin and IgG are present in the patient samples. 

Tuccito, N., Giamblanco, N., Licciardello, A., Marietta, G. (2007) Patterning of lactoferrin using functional SAMs of iron complexes. Chem. Commun., 2621-2623. 

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