STRUCTURAL EVIDENCE FOR A PH-SENSITIVE DILYSINE TRIGGER IN THE HEN OVOTRANSFERRIN N-LOBE - IMPLICATIONS FOR TRANSFERRIN IRON RELEASE

Members of the transferrin family of proteins are involved in Fe3+ tra nsport (serum transferrins) and are also believed to possess antimicro bial activity (ovotransferrins and lactoferrins). The structure of the monoferric N-terminal half-molecule of hen ovotransferrin, reported h ere at 2.3-angstrom resolution, reveals an unusual interdomain interac tion formed between the side-chain NZ atoms of Lys 209 and Lys 301, wh ich are 2.3 angstrom apart. This strong interaction appears to be an e xample of a low-barrier hydrogen bond between the two lysine NZ atoms, both of which are also involved in a hydrogen-bonding interaction wit h the aromatic ring of a tyrosine residue. Crystals of the protein wer e grown at pH 5.9, which is well below the usual pK(a) approximately 1 0 for a lysine side chain. We suggest that the pK(a) of either one or both of these residues lies below the pH of the structure determinatio n and is, therefore, not positively charged. This finding may serve to explain, on a molecular basis, the pH dependence of transferrin Fe3release. We propose that uptake of the Fe3+-transferrin complex into a n acidic endosome (viz., pH approximately 5.0) via receptor-mediated e ndocytosis will result in the protonation of both lysine residues. The close proximity of the two resulting positive charges, and their loca tion on opposite domains of the N-lobe, might well be the driving forc e that opens the two domains of the protein, exposing the Fe3+ ion and facilitating its release. Examination of amino acid sequences of othe r transferrins indicates that similar pH-sensitive dilysine triggers a re possible in the N-lobe, but not in the C-lobe, of most serum transf errins although such interactions have not been reported. Dilysine tri ggers are not possible in the C-lobe of hen ovotransferrin or in eithe r lobe of most lactoferrins.