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 β-strands in enhancing β-sheet stability has been examined quantitatively by NMR using rationally designed synthetic peptides [β-hairpin ( 2 β) and related 24-residue three-stranded antiparallel β-sheet ( 3 β)] which are significantly folded in aqueous solution. The two hairpin components of 3 β show quite different temperature-dependent stability profiles showing that a two-state model for folding (random coil ↔ three-stranded antiparallel β-sheet) is inappropriate. A four-state model for folding, involving intermediate C- and N-terminal β-hairpin conformations, is more consistent with the data. Thermodynamic analysis shows that folding of the C-terminal hairpin of 3 β is entropy-driven, as previously described for the isolated hairpin 2 β, 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 π−π 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 β-sheet. Both hairpins are associated with a significant Δ C p ° for folding, suggesting the burial of hydrophobic surface area as an important contributor to stability. We demonstrate quantitatively, by comparison of data for 2 β versus 3 β, that the folded population of the C-terminal β-hairpin is cooperatively enhanced by the interaction of the third strand.