title: Important residue (G46) in erythroid spectrin tetramer formation.
abstract: Spectrin tetramerization is important for the erythrocyte to maintain its unique shape, elasticity and deformability. We used recombinant model proteins to show the importance of one residue (G46) in the erythroid alpha-spectrin junction region that affects spectrin tetramer formation. The G46 residue in the erythroid spectrin N-terminal junction region is the only residue that differs from that in non-erythroid spectrin. The corresponding residue is R37. We believe that this difference may be, at least in part, responsible for the 15-fold difference in the equilibrium constants of erythroid and non-erythroid tetramer formation. In this study, we replaced the Gly residue with Ala, Arg or Glu residues in an erythroid alpha-spectrin model protein to give G46A, G46R or G46E, respectively. We found that their association affinities with a beta-spectrin model protein were quite different from each other. G46R exhibited a 10-fold increase and G46E exhibited a 16-fold decrease, whereas G46A showed little difference, when compared with the wild type. The thermal and urea denaturation experiments showed insignificant structural change in G46R. Thus, the differences in affinity were due to differences in local, specific interactions, rather than conformational differences in these variants. An intra-helical salt bridge in G46R may stabilize the partial domain single helix in alpha-spectrin, Helix C', to allow a more stable helical bundling in the alphabeta complex in spectrin tetramers. These results not only showed the importance of residue G46 in erythroid alpha-spectrin, but also provided insights toward the differences in association affinity between erythroid and non-erythroid spectrin to form spectrin tetramers.
authors.affiliation: Department of Chemistry, University of Illinois at Chicago, 845 W. Taylor Street, MC 111, Chicago, IL 60607, USA.
authors.forename: Jianxia
authors.initials: J
authors.lastname: Kang
authors.forename: Yuanli
authors.initials: Y
authors.lastname: Song
authors.forename: Akin
authors.initials: A
authors.lastname: Sevinc
authors.forename: Leslie W-M
authors.initials: LW
authors.lastname: Fung
mesh_terms: D019943:Amino Acid Substitution; D004912:Erythrocytes / Q000737:chemistry*; D005998:Glycine / Q000737:chemistry*; D016297:Mutagenesis, Site-Directed; D011489:Protein Denaturation; D055503:Protein Multimerization; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013049:Spectrin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D011994:Recombinant Proteins; D013049:Spectrin; D005998:Glycine