Dissecting the stability of a β-hairpin peptide that folds in water: NMR and molecular dynamics analysis of the β-turn and β-strand contributions to folding

NMR studies of the folding and conformational properties of a β-hairpin peptide, several peptide fragments of the hairpin, and sequence-modified analogues, have enabled the various contributions to β-hairpin stability in water to be dissected. Temperature and pH-induced unfolding studies indicate that the folding-unfolding equilibrium approximates to a two-state model. The hairpin is highly resistant to denaturation and is still significantly folded in 7 M urea at 298 K. Thermodynamic analysis shows the hairpin to fold in water with a significant change in heat capacity, however, ΔC(p)°in 7 M urea is reduced. V/Y→A mutations on one strand of the hairpin reduce folding to <10%, consistent with a hydrophobic stabilisation model. We show that in a truncated peptide (residues 6-16) lacking the hydrophobic residues on one β-strand, the type I' Asn-Gly turn in the sequence SINGKK is significantly populated in water in the absence of interstrand hydrophobic contacts. Unrestrained molecular dynamics simulations of unfolding, using an explicit solvation model, show that the conformation of the NG turn persists for longer than the AG analogue, which has a much lower propensity for type I' turn formation from a data base analysis of preferred turns. The origin of the high stability of the Asn-Gly turn is not entirely clear; data base analysis of 66 NG turns, together with molecular dynamics simulations, reveals no participation of the Asn side-chain in turn-stabilising interactions with the peptide backbone. However, hydration analysis of the molecular dynamics simulations reveals a pocket of 'high density' water bridging between the Asn side-chain and peptide main-chain that suggests solvent-mediated interactions may play an important role in modulating φ, ψ propensities in the NG turn region.