Diproline templates as folding nuclei in designed peptides. Conformational analysis of synthetic peptide helices containing amino terminal pro-pro segments

Abstract

The effect of N-terminal diproline segments in nucleating helical folding in designed peptides has been studied in two model sequences Piv-Pro-Pro-Aib-Leu-Aib-Phe-OMe (1) and Boc-Aib-Pro-Pro-Aib-Val-Ala-Phe-OMe (2). The structure of 1 in crystals, determined by X-ray diffraction, reveals a helical (α_R) conformation for the segment residues 2 to 5, stabilized by one 4→1 hydrogen bond and two 5→1 interactions. The N-terminus residue, Pro(1) adopts a polyproline II (P_II) conformation. NMR studies in three different solvent systems support a conformation similar to that observed in crystals. In the apolar solvent CDCl₃, NOE data favor the population of both completely helical and partially unfolded structures. In the former, the Pro-Pro segment adopts an α_R-α_R conformation, whereas in the latter, a P_II-α_R structure is established. The conformational equilibrium shifts in favor of the P_II-α_R structure in solvents like methanol and DMSO. A significant population of the Pro(1)-Pro(2) cis conformer is also observed. The NMR results are consistent with the population of at least three conformational states about Pro-Pro segment: trans α_R-α_R, trans P_II-α_R and cis P_II-α_R. Of these, the two trans conformers are in rapid dynamic exchange on the NMR time scale, whereas the interconversion between cis and trans form is slow. Similar results are obtained with peptide 2. Analysis of 462 diproline segments in protein crystal structures reveals 25 examples of the α_R-α_R conformation followed by a helix. Modeling and energy minimization studies suggest that both P_II-α_R and α_R-α_R conformations have very similar energies in the model hexapeptide 1.