Evidence for the functional equivalence of the iron-binding sites of rat transferrin.
The role of the two iron-binding sites of rat transferrin in the exchange of iron with cells has been assessed using urea polyacrylamide gel electrophoresis to separate and quantitate the four possible molecular species of transferrin generated during the incubation of 125I-labelled transferrin with rat reticulocytes and hepatocytes. Addition of diferric transferrin to reticulocytes led directly to the appearance of apotransferrin together with small and comparable amounts of the two monoferric transferrins. After 2 h 44.8% of the iron had been removed by the cells, and of the iron-depleted transferrin 71.8% was apotransferrin, the remainder being monoferric transferrin, 16.1% with N-terminal iron and 12.1% with C-terminal iron. A similar pattern emerged with hepatocytes, but the rate of iron removal was slower and the proportion of apotransferrin generated was lower. After 4 h 10.9% of the iron had been removed from the transferrin and the distribution of the iron-depleted protein was: apotransferrin 26.9% and monoferric (N-terminal) 39.2%, (C-terminal) 33.9%. The appearance of apotransferrin during each incubation and the generation of both monoferric transferrins suggest that both cell types are able to remove iron from differic transferrin in pairwise fashion and that they do not appreciably distinguish between the two iron-binding sites of the protein. Release of iron from hepatocytes to apotransferrin lead to the appearance of both monoferric species and then to increasing amounts of diferric transferrin. The process of iron release did not seem to distinguish between the vacant iron-binding sites of transferrin.
D000818:Animals; D001665:Binding Sites; D007501:Iron / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008297:Male; D011485:Protein Binding; D051381:Rats; D011919:Rats, Inbred Strains; D012156:Reticulocytes / Q000378:metabolism*; D014168:Transferrin / Q000378:metabolism*
D014168:Transferrin; D007501:Iron