A mechanism for iron uptake by transferrin.

Iron uptake by transferrin from iron nitrilotriacetate (FeNAc3) in the presence of bicarbonate has been investigated in the pH range 6.5-8. Apotransferrin, in interaction with bicarbonate, extracts iron from FeNAc3, without the formation of an intermediate protein-iron-ligand mixed complex (iron-exchange-equilibrium constant, K1=1 +/- 0.05; direct second-order-rate constant, k1=8.0x10(4) +/- 0.5x10(4)M(-1)s(-1)., reverse second-order-rate constant, k-1=7.5x10(4) +/- 0.5x10(4)M(-1)s(-1). The newly formed iron-protein complex loses a single proton (proton-dissociation constant, Ka=16 +/- 1.5nM) and then undergoes a modification of its conformation followed by loss of two or three protons (first-order-rate constant, K2=2.80 +/- 0.10s-1). This includes a new modification in the conformation (first-order-rate constant, K2=6.2x10(2) +/- 0.3x10(-2)s(-1). This second modification in conformation controls the rate of iron uptake by the N-site of the protein and is followed by loss of one proton (K3a=6.80 nM). Finally, the holoprotein or the monoferric transferrin in its final equilibrated state is produced by a third modification in the conformation that occurs after approximately 3000 s. Iron uptake by the N-site does not occur when the apotransferrin interacts with bicarbonate. Nevertheless, it occurs with the monoferric transferrin, in which iron is bound to the C-site, in its final state of equilibrium by a mechanism similar to that of iron uptake by the C-site of apotransferrin. These modifications in the conformation of the protein occur after iron uptake by the C-site and may be important for the recognition of the protein by its receptor prior to iron delivery by endocytosis.