title: Mapping the binding sites of human erythrocyte ankyrin for the anion exchanger and spectrin.
abstract: This report describes initial characterization of the binding sites of ankyrin for spectrin and the anion exchanger using defined subfragments isolated from purified ankyrin domains. The spectrin-binding domain of ankyrin is comprised of two subdomains: an acidic, proline-rich region (pI = 4) involving the amino-terminal 80 residues from 828 to 908 and a basic region (pI = 8.8) that extends from 898 to 1386. The amino-terminal 70 amino acids of the spectrin-binding domain are critical for association with spectrin, since a subfragment missing this region is only 5% as active as the intact domain in displacing binding of spectrin to inside-out membrane vesicles, while deletion of the first 38 residues of the acidic domain results in a 10-fold reduction in activity. The anion exchanger-binding site is confined to an 89-kDa domain that was isolated and characterized as a globular molecule with approximately 30% alpha-helical configuration. A subfragment of the 89-kDa domain extending from residues 403 to 779 (or possibly 740) retains ability to associate with the anion exchanger. The 89-kDa domain is comprised of a series of tandem repeats of 33 amino acids that extend from residues 35 to 778 (Lux, S., John, K., and Bennett, V. (1990) Nature 344, 36-42). The activity of residues 403-779 demonstrates that the 33-amino acid repeats of the 89-kDa domain are responsible for association between ankyrin and the anion exchanger. The 33-amino acid repeating sequence of ankyrin represents an ancient motif also found in proteins of Drosophila, yeast, and Caenor habditis elegans. The finding that the 33-amino acid repeating sequence is involved in interaction with the anion exchanger implies that this motif may perform a role in molecular recognition in diverse proteins.
authors: Howard Hughes Medical Institute, Duke University Medical Center, Durham, North Carolina 27710.
authors: L H
authors: LH
authors: Davis
authors: V
authors: V
authors: Bennett
mesh_terms: D000595:Amino Acid Sequence; D027321:Anion Transport Proteins; D017487:Ankyrins; D001665:Binding Sites; D001798:Blood Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D002352:Carrier Proteins / Q000378:metabolism*; D002942:Circular Dichroism; D004910:Erythrocyte Membrane / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008565:Membrane Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011487:Protein Conformation; D013049:Spectrin / Q000378:metabolism*
chemical_list: D027321:Anion Transport Proteins; D017487:Ankyrins; D001798:Blood Proteins; D002352:Carrier Proteins; D008565:Membrane Proteins; D013049:Spectrin