Minimalist protein design:: A β-hairpin peptide that binds ssDNA

A 28-residue β-hairpin dimer (WKWK)2 with two Trp and two Lys residues on one face of each β-sheet was shown to form a complex with single-stranded oligonucleotides at low micromolar concentrations. Each β-hairpin of the dimer contains a cross-strand Trp-Trp pair in a diagonal orientation which has previously been shown to create a cleft for the intercalation of aromatic guests such as adenine (J. Am. Chem. Soc. 2003, 125, 9580). The β-hairpin dimer binds 5-residue ssDNA sequences 5-AAAAA-3-, 5-TTTTT-3-, and 5-CCCCC-3- in water with dissociation constants in the range of 12-30 μM. A weak energetic preference for binding to sequence 5-AAAAA-3- was observed, which is believed to result from stronger stacking interactions between Trp and the adenine base. The interaction of 5-AAAAA-3- with the Lys and Trp residues of the peptide was evident by NMR, and a 1:1 association was demonstrated. The recognition of an 11-residue ssDNA sequence occurred with a dissociation constant of 3 μM under near-physiological ionic strength and pH, demonstrating that the β-hairpin dimer binds ssDNA as strongly as many naturally occurring proteins. The salt dependence of the interaction of the 11-residue oligonucleotide with the peptide dimer indicates that Trp-nucleobase stacking interactions contribute about -4 kcal/mol to recognition, which is much greater than the contribution of nonionic interactions in unstructured peptides containing Trp. Moreover, recognition of the ssDNA demonstrated reduced salt dependence relative to the corresponding duplex, resulting in selectivity for ssDNA under high salt conditions. Peptide (WKWK)2 is a relevant mimic of OB-fold (oligonucleotide/oligosaccharide-binding) proteins which bind ssDNA on the surface of a β-sheet. Copyright © 2005 American Chemical Society.