One of the puzzles of iron metabolism is the mechanism by which cells are so effective at removing ferric ion from the powerful iron chelating protein transferrin. The mechanism of iron release from transferrin is also highly relevant to the development of a more effective drug for the treatment of iron overload which afflicts victims of Beta-thalassemia and sickle cell anemia. Iron release to low molecular weight ligands has been studied for several types of ligands, including polyphosphates, hydroxamates, amino acids, and catechols, and conflicting mechanisms have been proposed. This project involves a systematic study of the kinetics of iron removal by a series of aminophosphonic acid and phenolic ligands. A major objective of this study is to distinguish between the two most prominent proposed mechanisms for iron removal. This involves a full evaluation of the reaction rate as a function of the concentration of both the competing ligand and bicarbonate, and the determination of the activation parameters for the rate constants for each ligand. In cases where saturation kinetics are observed, the hyperbolic dependence of the rate of iron removal on the concentration of the ligand will be established, and pseudo first order rate constants will be determined for the rate limiting step at high ligand concentrations. Rate constants will also be determined for iron removal from both N-terminal and C-terminal mono(ferric)transferrin. Spectra will also be recorded for mixtures of ferric-transferrin with monodentate ligands such as phosphate, phenol, and acetate to characterize any mixed-ligand complex which may form under conditions where no significant amount of iron removal will occur.
