Figure 1.

Native and recombinant proteins used in this study. (A) Coomassie blue-stained Laemmli SDS-PAGE. The assembled and purified minispectrins are in lanes 1 and 2. (B) CD spectra of the recombinant α18-21EF (EF), α18-21EFΔ13 (EFΔ13), and β1-4ABD peptides. The spectra display the troughs at 208 nm and 222 nm characteristic of α-helical peptides, indicating that the recombinant proteins are folded. Structure estimation predicts approximately 75% α-helix for the βABD1-4 peptide, approximately 80% for α18-21EF, and approximately 85% for α18-21EFΔ13. However, the latter estimate is rough because the error in the estimate is high. Very little β-structure (< 2%) is predicted. The data approximate what would be expected from the known structures of the highly α-helical spectrin repeats and EF hands that constitute most of the α18-21EF peptide and the spectrin repeats and CH domains that make up much of the βABD1-4 peptide. The fact that the mutant peptide α18-21EFΔ13 appears to be even more helical than α18-21EF suggests that the C-terminal 13 amino acid deletion substantially alters the structure of the EF domain.