Simeon Pollack

Iron removal from transferrin. An experimental study.

We have studied the facilitation of iron transfer from transferrin to desferrioxamine by various anions. Most of the anions which can substitute for HCO-3 in the ternary complex of transferrin - Fe - HCO3 do not facilitate iron transfer; anions which do facilitate iron transfer do not necessarily form stable ternary complexes. Combinations of anions effective in transfer have a less-than-additive effect, suggesting a common reaction pathway. We suggest that the transfer of iron from transferrin to desferrioxamine involves a substitution step and a subsequent chelation step, and that the efficiency of the overall reaction is a function of both these attributes of the anion.

Chelate mediated transfer of iron from transferrin to desferrioxamine.

SUMMARY. Desferrioxamine, widely used for the treatment of iron overload in Cooleys anaemia, binds iron so tightly that it should quantitatively remove iron from transferrin. Studies conducted in vivo and in vitro, however, have failed to demonstrate significant depletion of transferrin‐bound iron by a stoichiometric excess of desferrioxamine. However, low molecular weight chelating agents, capable of forming ternary complexes with transferrin and ferric iron, can promote a rapid transfer of iron from transferrin to desferrioxamine. A possible mechanism for this facilitated exchange is offered.

Desferrioxamine Suppresses Plasmodium falciparum in Aotus Monkeys

Abstract Clinical observation has suggested that iron deficiency may be protective in malaria, and we have found that desferrioxamine (DF), an iron-specific chelating agent, inhibited Plasmodium falciparum growth in vitro. It was difficult to be confident that DF would be effective in an intact animal, however, because continuous exposure to DF was required in vitro and, in vivo, DF is rapidly excreted. Also, the in vitro effect of DF was overcome by addition of iron to the culture and in vivo there are potentially high local iron concentrations when iron is absorbed from the diet or released from reticuloendothelial cells. We now show that DF given by constant subcutaneous infusion does suppress parasitemia in P. falciparum-infected Aotus monkeys.

Plasmodium falciparum takes up iron from transferrin

Plasmodium falciparum, cultured in vitro, is inhibited by desferrioxamine, a specific iron chelating agent, despite the abundant haem iron which surrounds the parasite. In this report we show that the intraerythrocytic parasite obtains iron from transferrin. The parasite may therefore be freed of a requirement to catabolize haem to obtain iron thus minimizing haem toxicity.

Immature red cells have ferritin receptors

Abstract Ferritin binds to immature red cells. The binding appears to be specific: (1) It is abolished by a large competing dose of nonradioactive ferritin. (2) There is little binding of ferritin to mature red cells. (3) Other high molecular weight proteins (gamma globulin and thyroglobulin) are not bound.

Inability to detect transferrin receptors on P. falciparum parasitized red cells.

Summary The mechanism by which P. falciparum takes up iron from transferrin has been explored. Binding of 125I labelled transferrin to parasitized red cells at 37°C is two‐fold greater than to control cells; at 0°C there is no significant difference. The binding is non‐specific as judged from the following: it is not saturable: it is not limited to transferrin as lactoferrin (which has iron binding domains) and bovine serum albumin (which does not) also bind in excess to parasitized red cells. A transferrin receptor complex could not be demonstrated when parasitized red cells, to which 125I transferrin was bound, were solubilized in Triton X100. Previous observation showed that uptake of transferrin iron by parasitized red cells is not accompanied by equimolar uptake of transferrin protein. We therefore suggest that non‐specifically bound transferrin is endocytosed, that the protein is degraded and the iron selectively retained.

Iron bound to low MW ligands: interactions with mitochondria and cytosolic proteins.

Abstract: The iron in the low MW pool of the cell is the precursor of iron in haem and is bound primarily to ATP. This precursor‐product relationship suggested that reticulocytes might accumulate ATP‐iron if their haem synthesis were blocked. However reticulocytes, treated with succinyl‐acetone or rotenone and taking up iron from transferrin, accumulated iron in nonhaem cytosolic proteins and in mitochondria and not in the low MW pool. This was demonstrated by NMR and also by disrupting the cell with shear stress, separating the cytosol and pellet and fractionating the cytosol with ammonium sulfate. This constancy of the low MW iron pool in the face of blocked haem synthesis could not be explained by saturation of cytosolic ATP or by sluggish exchange of the low MW pool with other compartments. Rather, nonhaem cytosolic proteins and mitochondria appeared to have a higher affinity for iron and to exchange it rapidly with that in the low MW pool.

Early events in guinea pig reticulocyte iron uptake

Hemolysates, prepared from guinea pig reticulocytes incubated with 59Fe-labelled serum, can be resolved into five peaks utilizing molecular sieve chromatography: ferritin, transferrin, hemoglobin, an Mx 17 000 fraction, and a low molecular weight fraction. The hemoglobin peak also contains a nonhemoglobin component (III-X), demonstrated by heme extraction and by isoelectric focusing. Transferrin, the III-X component and the low molecular weight fraction are the first to accumulate radioactive iron during the reticulocyte incubation. The 59Fe in each of these also chases. Therefore, a role for these components as precursors to iron incorporation into heme is suggested.

Guinea pig intestinal iron binding protein.

Abstract An iron binding protein, isolated from guinea pig intestinal mucosa, was compared to guinea pig transferrin. Both had a molecular weight of approximately 80,000. The intestinal iron-binding protein consisted of 2 subunits of equal molecular weight; transferrin had no subunits. Transferrin showed an absorbance peak at 470 nm; the intestinal iron-binding protein had no visible absorbance but did have a peak at 336 nm. Electron spin resonance spectra of the two proteins dfffered. Significant differences on amino acid analysis were also identified.

P. Falciparum infected erythrocytes are capable of endocytosis

SummaryP. falciparum, an intraerythrocytic parasite, obtains nourishment primarily through phagocytosis of the host cytosol but also through the incorporation of extracellular small molecules which enter through the parasitized red cells membrane via pores. Normal mature erythrocytes are incapable of endocytosis. Several lines of evidence suggest that extracellular large molecules may be taken up when the mature red cell is parasitized byP. falciparum, but direct evidence has been lacking. We now report the use of ferritin, an electron dense protein, to demonstrate endocytosis inP. falciparum infected red cells.Parasitized red cells incubated with ferritin internalize that macromolecule as demonstrated by electron microscopy. While normal red cells incubated with ferritin took up none of the tracer molecule, parasitized red cells internalized substantial amounts. In addition both ferritin and apoferritin inhibited the growth ofP. falciparum in a dose dependent fashion, again indicating endocytosis of a macromolecule. These data indicate thatP. falciparum can somehow stimulate the mature erythrocyte to engage in endocytosis. We also note that both infected and non-infected red cells in a culture in whichP. falciparum is growing become abnormally sticky for ferritin. Moreover, parasitized red cells bind I125-transferrin while non-parasitized erythrocytes do not. These observations suggest that a soluble parasite product alters the red cell membrane in a non-global manner, causing selective effects in relation to different proteins.