Structure, folding, and energetics of cooperative interactions between the β-strands of a de novo designed three-stranded antiparallel β-sheet peptide

The effect of cooperative interactions between beta-strands in enhancing beta-sheet stability has been examined quantitatively by NMR using rationally designed synthetic peptides [beta-hairpin (2 beta) and related 24-residue three-stranded antiparallel beta-sheet (3 beta)] which are significantly folded in aqueous solution. The two hairpin components of 3 beta show quite different temperature-dependent stability profiles showing that a two-state model for folding (random coil <-> three-stranded antiparallel beta-sheet) is inappropriate. A four-state model for folding, involving intermediate C- and N-terminal beta-hairpin conformations, is more consistent with the data. Thermodynamic analysis shows that folding of the C-terminal hairpin of 3 beta is entropy-driven, as previously described for the isolated hairpin 2 beta, but that the N-terminal hairpin, which is stabilized by a motif of aromatic residues (W4, F6, and Y11), is enthalpy-driven, consistent with stabilization through pi-pi interactions that are electrostatic in origin. NOE data, as well as structure calculations, support the formation of this stabilizing motif on one face of the beta-sheet. Both hairpins are associated with a significant Delta C(p)degrees for folding, suggesting the burial of hydrophobic surface area as an important contributor to stability. We demonstrate quantitatively, by comparison of data for 2 beta versus 3 beta, that the folded population of the C-terminal beta-hairpin is cooperatively enhanced by the interaction of the third strand.