Transferrin active sites

Since malonate (Structure I) is able to fulfill the role of the anion in transferrin, it seemed reasonable to see whether spin-labeled derivatives of malonate could serve as probes of the active sites. Two such spin-labled derivatives were prepared and tentatively identified as having structures II (N-4-(2,2,6,6-tetramethylpiperidin-l-oxyl)malonamide) and III (N-4-(2,2,6,6-tetramethylpiperidin-l-oxyl)malonate). Similar results were obtained with each (Figure 3). Upon mixing Fe(III), transferrin, and II at low pH, and then raising the pH to near-neutrality with C02-free ammonia, the characteristic orange-red color of the ternary Fe-transferrin-anion complex is promptly displayed. However, the anticipated EPR signal of the nitroxide spin-label is not observed, presumably because it is broadened beyond detectability by its proximity... 

Vanadium(IV) complexes with two dipeptides glycyl-tyrosine and glycyl-phenylalanine (Gly-Tyr and Gly-Phe, respectively) and their oxovanadium(lV) and (V) complexes have been observed.382, 01 The complex formed with Gly-Tyr showed coordination by the peptide backbone moieties, and no interaction from the distal phenolic hydroxy group.601 A histidine-tyrosine derivatized peptide was shown to complex Viv in a pentadentate manner (120).564 The adduct between VlV and a model of the active-site peptide of protein tyrosine phosphatases has been spectroscopically characterized.602 Similar characterization of in serum suggests that protein complexes form, and that both Viv and Vv can exist as complexes with transferrin and albumin.603 ESEEM has been used to characterize the complexes of apoferritin with V02+ and suggests that the carboxylates, one water molecule, and one histidine ligand fill the coordination sites of the vanadium.604... 

When transferrin is labeled with FITC, the coordinated iron atom at the active site acts as an effective quencher of emitted light, so that this prepared substrate exhibits low emission intensity at 525 nm. Upon exposure to protease, cleavage of the polypeptide occurs, resulting in the release of FITC labeled amino acids and oligopeptides. Because the fluorophore and the quencher are no longer fixed in... 

Unlike the sharp signals that are observed for Cr(V), those of Cr(III) are typically very broad and difficult to observe at room temperature. The signals are typically centered around g values of 1.98-2.00 (13, 43). While EPR spectroscopy is very useful for the detection of Cr(III) products from redox reactions of Cr in higher oxidation states (44), it is of limited value in determining the namre of the donor atoms bound to the metal ion. Nonetheless, it is still an important technique for study of the bioinorganic chemistry of Cr(lll). One application includes smdies on frozen solutions (77 K) of the active site for binding of Cr(lll) in transferrin (45), which is believed to be the major transport protein for Cr(lll) in mammals (Section IV.B). 

Li, B., Schopfer, L.M., Grigoryan, H., et al., 2009a. Tyrosines of human and mouse transferrin covalently labeled by organophosphorus agents a new motif for binding to proteins that have no active site serine. Toxicol. Sci. 107 (1), 144-155. 

Lactoferrin resembles transferrin in terms of molecular weight, amino-acid sequence homology and number of Fe(lII) binding sites. Lactoferrin is released from activated PMNs upon degranulation and may play a role in myelopoiesis, primary antibody response, lymphocyte proliferation, cytokine production and complement activation. 

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