First step toward the quantitative identification of peptide 3₁₀-helix conformation with NMR spectroscopy: NMR and x-ray diffraction structural analysis of a fully-developed 3₁₀-helical peptide standard

We have synthesized by solution methods and fully characterized the Nα-blocked heptapeptide methylamide mBrBz-[L-Iva-L-(αMe)Val]₂- L-(αMe)Phe-L-(αMe)Val-L-Iva-NHMe, fully based on conformationally constrained Cα-methylated α-amino acids. An X-ray diffraction investigation of the Nα-benzyloxycarbonylated analogue showed that in the crystal state both independent molecules (A and B) in the asymmetric unit of the peptide adopt a fully developed, regular, right- handed 3₁₀-helical structure, although molecule A would be slightly distorted at the C-terminal residue. Solution conformational analysis on the mBrBz-blocked peptide was carried out in CDCl₃ by means of NMR spectroscopy. For structure determination we performed restrained molecular dynamics simulations in CDCl₃ based on a search of the conformational space derived from a simulated annealing strategy. For this peptide the NMR observables can be described by a single backbone conformation, more specifically a rigid 3₁₀-helix spanning the amino acid sequence from residue 1 to residue 6. The C-terminal methylamido NH group seems to be involved simultaneously in two H-bonds (with the preceding i - 3 and i - 4 carbonyl groups). Although in this peptide model there are no distinct NOE distances for discriminating 3₁₀- versus α-helix conformation, the sum of all NMR-derived restraints clearly results in a 3₁₀-helical structure. Convergence from different starting structures (including an α-helix) into a 3₁₀- helix was observed.