title: Studies of the erythrocyte spectrin tetramerization region.
abstract: Human erythrocyte spectrin dimers associate at the N-terminal region of alpha spectrin (alpha N) and the C-terminal region of beta-spectrin (beta C) to form tetramers. We have prepared model peptides to study the tetramerization region. Based on phasing information obtained from enzyme digests, we prepared spectrin fragments consisting of the first 156 amino-acid residues and the first 368 amino-acid residues of alpha-spectrin (Sp alpha 1-156 and Sp alpha 1-368, respectively), and found that both peptides associate with a beta-spectrin model peptide, with an affinity similar to that found in alpha beta dimer tetramerization. Spin label EPR studies show that the region consisting of residues 21-46 in alpha-spectrin is helical even in the absence of its beta-partner. Multi-dimensional nuclear magnetic resonance studies of samples with and without a spin label attached to residue 154 show that Sp alpha 1-156 consists of four helices, with the first helix unassociated with the remaining three helices, which bundle to form a triple helical coiled coil bundle. A comparison of the structures of erythrocyte spectrin with other published structures of Drosophila and chicken brain spectrin is discussed. Circular dichroism studies show that the lone helix in Sp alpha-156 associates with helices in the beta peptide to form a coiled coil bundle. Based on NMR and CD results, we suggest that the helices in Sp alpha 1-156 exhibit a looser (frayed) conformation, and that the helices convert to a tighter conformation upon association with its beta-partner. This suggestion does not rule out possible conversion of a non-structured conformation to a structured conformation in various parts of the molecule upon association. Spectrin mutations at residues 28 and 45 of alpha-spectrin have been found in patients with hereditary elliptocytosis. NMR studies were also carried out on Sp alpha 1-156R28S, Sp alpha 1-156R45S and Sp alpha 1-156R45T. A comparison of the structures of Sp alpha 1-156 and Sp alpha 1-156R28S, Sp alpha 1-156R45S and Sp alpha 1-156R45T is discussed.
authors.affiliation: Center for Pharmaceutical Biotechnology, University of Illinois at Chicago, Chicago, IL 60607, USA.
authors.forename: S
authors.initials: S
authors.lastname: Park
authors.forename: S
authors.initials: S
authors.lastname: Mehboob
authors.forename: B H
authors.initials: BH
authors.lastname: Luo
authors.forename: M
authors.initials: M
authors.lastname: Hurtuk
authors.forename: M E
authors.initials: ME
authors.lastname: Johnson
authors.forename: L W
authors.initials: LW
authors.lastname: Fung
mesh_terms: D000595:Amino Acid Sequence; D001665:Binding Sites; D004578:Electron Spin Resonance Spectroscopy; D004912:Erythrocytes / Q000378:metabolism*; D004926:Escherichia coli; D006801:Humans; D019906:Nuclear Magnetic Resonance, Biomolecular; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D013049:Spectrin / Q000737:chemistry* / Q000378:metabolism
chemical_list: D010446:Peptide Fragments; D021122:Protein Subunits; D011994:Recombinant Proteins; D013049:Spectrin