The three-dimensional structure of the anti-apoptotic protein Bcl-xL complexed to a 25-residue peptide from the death promoting region of Bad was determined using NMR spectroscopy. Although the overall structure is similar to Bcl-xL bound to a 16-residue peptide from the Bak protein (Sattler et al., 1997), the Bad peptide forms additional interactions with Bcl-xL. However, based upon site-directed mutagenesis experiments, these additional contacts do not account for the increased affinity of the Bad 25-mer for Bcl-xL compared to the Bad 16-mer. Rather, the increased helix propensity of the Bad 25-mer is primarily responsible for its greater affinity for Bcl-xL. Based on this observation, a pair of 16-residue peptides were designed and synthesized that were predicted to have a high helix propensity while maintaining the interactions important for complexation with Bcl-xL. Both peptides showed an increase in helix propensity compared to the wild-type and exhibited an enhanced affinity for Bcl-xL.

Six helix surface positions of protein G (Gβ1) were redesigned using a computational protein design algorithm, resulting in the five fold mutant Gβ1m2. Gβ1m2 is well folded with a circular dichroism spectrum nearly identical to that of Gβ1, and a melting temperature of 91 °C, ∼6 °C higher than that of Gβ1. The crystal structure of Gβ1m2 was solved to 2.0 Å resolution by molecular replacement. The absence of hydrogen bond or salt bridge interactions between the designed residues in Gβ1m2 suggests that the increased stability of Gβ1m2 is due to increased helix propensity and more favorable helix dipole interactions.