Unique stabilizing interactions identified in the two‐stranded α‐helical coiled‐coil: Crystal structure of a cortexillin I/GCN4 hybrid coiled‐coil peptide

Abstract

We determined the 1.17 Å resolution X‐ray crystal structure of a hybrid peptide based on sequences from coiled‐coil regions of the proteins GCN4 and cortexillin I. The peptide forms a parallel homodimeric coiled‐coil, with C α backbone geometry similar to GCN4 (rmsd value 0.71 Å). Three stabilizing interactions have been identified: a unique hydrogen bonding–electrostatic network not previously observed in coiled‐coils, and two other hydrophobic interactions involving leucine residues at positions e and g from both g‐a' and d‐e' interchain interactions with the hydrophobic core. This is also the first report of the quantitative significance of these interactions. The GCN4/cortexillin hybrid surprisingly has two interchain Glu‐Lys' ion pairs that form a hydrogen bonding network with the Asn residues in the core. This network, which was not observed for the reversed Lys‐Glu' pair in GCN4, increases the combined stability contribution of each Glu‐Lys' salt bridge across the central Asn15‐Asn15′ core to ∼0.7 kcal/mole, compared to ∼0.4 kcal mole −1 from a Glu‐Lys' salt bridge on its own. In addition to electrostatic and hydrogen bonding stabilization of the coiled‐coil, individual leucine residues at positions e and g in the hybrid peptide also contribute to stability by 0.7 kcal/mole relative to alanine. These interactions are of critical importance to understanding the stability requirements for coiled‐coil folding and in modulating the stability of de novo designed macromolecules containing this motif.