title: Characterization and structural analysis of a functional human serum transferrin variant and implications for receptor recognition.
abstract: The nucleotide and amino acid substitutions leading to the only known functional variant of human serum transferrin have been characterized by sequencing of a peptide produced by cyanogen bromide digestion and genomic PCR coupled with cycle sequencing, respectively. There is an amino acid substitution at position 394 (Gly-->Arg) resulting from a mutational transition, G-->A, in the first nucleotide of the codon GGG. The Zn(2+)-, Al(3+)-, and Cu(2+)-binding properties of the variant, ascertained by UV difference spectra and, in the case of copper, protein fluorescence quenching, confirm that these metals binds to only one of the two sites. Solution X-ray scattering measurements indicate that the lobe (the C-lobe) containing the mutation remains "open" in the iron-bound state, and modeling studies suggest that this is a consequence of the formation of a salt bridge between Arg394 in the variant protein and Asp392, one of the iron-binding ligands in the C-lobe. This rationalizes for the first time the observed reduction in receptor affinity of the diferric variant protein for PHA-stimulated lymphocytes [Young, S.P., et al. (1984) Br. J. Haematol. 56, 581-587] and here repeated with K562 cells. These data lend support to the hypothesis that the closed conformation for both lobes contributes to receptor recognition.
authors.affiliation: Division of Biochemistry & Molecular Biology, UMDS, Guy's Hospital, London, U.K.
authors.forename: R W
authors.initials: RW
authors.lastname: Evans
authors.forename: J B
authors.initials: JB
authors.lastname: Crawley
authors.forename: R C
authors.initials: RC
authors.lastname: Garratt
authors.forename: J G
authors.initials: JG
authors.lastname: Grossmann
authors.forename: M
authors.initials: M
authors.lastname: Neu
authors.forename: A
authors.initials: A
authors.lastname: Aitken
authors.forename: K J
authors.initials: KJ
authors.lastname: Patel
authors.forename: A
authors.initials: A
authors.lastname: Meilak
authors.forename: C
authors.initials: C
authors.lastname: Wong
authors.forename: J
authors.initials: J
authors.lastname: Singh
mesh_terms: D000535:Aluminum / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D003062:Codon; D003300:Copper / Q000378:metabolism; D003488:Cyanogen Bromide; D006801:Humans; D008958:Models, Molecular; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009154:Mutation; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D011990:Receptors, Transferrin / Q000378:metabolism*; D012542:Scattering, Radiation; D017421:Sequence Analysis; D013056:Spectrophotometry, Ultraviolet; D014168:Transferrin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D014965:X-Rays; D015032:Zinc / Q000378:metabolism
chemical_list: D003062:Codon; D011990:Receptors, Transferrin; D014168:Transferrin; D003300:Copper; D000535:Aluminum; D015032:Zinc; D003488:Cyanogen Bromide