TRANSFERRINS - A MECHANISM FOR IRON UPTAKE BY LACTOFERRIN

Iron uptake by bovine lactoferrin from nitrilotriacetatoFe(III) [FeN(A c)(3)] in the presence of bicarbonate has been investigated at pH 7.1- 8.7. Deprotonated apolactoferrin interacting with bicarbonate or carbo nate extracts iron from nitrilotriacetatoFe(III); the direct second-or der rate constant k(1) = (4.90 +/- 0.20) x 10(4) M-1 s(-1), a reverse second-order rate constant k(-1) = (1.80 +/- 0.05) x 10(5) M-1 s(-1), and the iron-exchange equilibrium constant K-1 = 0.25 +/- 0.05. The ne wly formed iron-protein complex loses a single proton with proton diss ociation constant K-3a = (17 +/- 0.5) nM, then undergoes a modificatio n in its conformation followed by the loss of two or three protons; th e first-order rate constant k, = (1.0 +/- 0.10) s(-1). This induces a new modification in the conformation; the first-order rate constant k( 3) = (8.75 +/- 0.40) x 10(-3) s(-1). This second modification in confo rmation controls the rate of iron uptake by the N site of the protein and is followed by a single proton loss; K-5a = 8.0 nM. Finally, the h oloprotein or the monoferric lactoferrin in their final equilibrated s tates are produced by a third modification in the conformation occurri ng in about 9000 s. The mechanism of iron uptake by lactoferrin is ver y similar to that of serum transferrin with a cooperativity between th e C and N sites upon iron uptake but with lower rates, higher affiniti es and at least one more proton loss involved. These differences may b e the result of slight discrepancies in the intimate structures of bin ding sites for serum transferrin and lactoferrin. In order to analyse the cooperativity between these iron-binding sites, the three-dimensio nal position of the chain of amino acid residues separating the N and C lobes of human apo-, holo-and dicopper-lactoferrin have been compare d by the recognition of the three-dimensional shape dissimilarity prog ram. The interlobe peptides of human hololactoferrin and apolactoferri n showed only 75.5% tridimensional similarity, indicating that iron up take affects the three-dimensional structure of the interlobe chain.