Identification of spectrin-RESA binding sites. (A) Schematic representation of RESA protein and recombinant fragments. The locations of the 5′ and 3′ repeat regions, and the positions in the sequence of the expressed fragments are indicated. (B) Schematic representation of the spectrin α- and β-chains, showing the domain structure, and locations in the sequence of expressed recombinant fragments. The boundaries of all spectrin fragments and single repeats were defined by SMART annotations. (C) Binding of spectrin dimer to RESA fragments. Spectrin dimer was incubated for 30 minutes at room temperature with each of the MBP-tagged RESA fragments, and binding was assessed by pull-down assay; the bound fragment was detected by blotting with antispectrin antibody after SDS-PAGE. (D) Binding of recombinant spectrin fragments to the RESA F5 fragment. Recombinant GST-tagged spectrin fragments were incubated with RESA F5 and binding was assayed as described for panel C, using anti-GST antibody for detection. (E) Binding of single GST-tagged β-spectrin repeats to RESA F5 fragment. Binding assays were performed as above. Note binding to βR16 only.