Alternative structural state of transferrin - The crystallographic analysis of iron-loaded but domain-opened ovotransferrin N-lobe

Transferrins bind Fe3+ very tightly in a closed interdomain cleft by the coordination of four protein ligands (Asp(60), Tyr(92) Tyr(191) and His(250) in ovotransferrin N-lobe) and of a synergistic anion, physiologically bidentate CO32-. Upon Fe3+ uptake, transferrins undergo a large scale conformational transition: the apo structure with an opening of the interdomain cleft is transformed into the closed hole structure, implying initial Fe3+ binding in the open form. To solve the Fe3+-loaded, domain-opened structure, an ovotransferrin N-lobe crystal that had been grown as the apo form was soaked with Fe3+ -nitrilotriacetate, and its structure was solved at 2.1 Angstrom resolution. The Fe3+-soaked form showed almost exactly the same overall open structure as the iron-free apo form. The electron density map unequivocally proved the presence of an iron atom with the coordination by the two protein ligands of Tyr(92)-OH and Tyr(191)-OH. Other Fe3+ coordination sites are occupied by a nitrilotriacetate anion, which is stabilized through the hydrogen bonds with the peptide NH groups of Ser(122), Ala(123), and Gly(124) and a side chain group of Thr(117). There is, however, no clear interaction between the nitrilotriacetate anion and the synergistic anion binding site, Arg(121).