Characterization of α-helix structures in polypeptides, revealed by 13CO⋯H– 15N hydrogen bond lengths determined by 13C REDOR NMR

Abstract

We have determined accurate 13C– 15N interatomic distances of mono-dispersed sequential oligopeptides, [1- 13C]Leu 8-, [ 15N]Ala 12-labeled (Phe-Leu-Ala) 6, and a variety of its derivatives in which Leu, at the residue11, was replaced by Ala, Phe, Gly or d-Ala, by means of rotational echo double resonance (REDOR) experiments. In order to minimize a plausible contribution from the peak intensities of natural abundant signals from amino-acid residues other than labeled, 13C NMR signals from such contributions were subtracted from the full echo amplitude of the doubly labeled spectra as reference. We have determined the interatomic distance as 4.5±0.1 Å for five kinds of sample examined without any significant deviation among them. Because [1- 13C]Leu 8 and [ 15N]Ala 12 form CO⋯HN hydrogen bond in the case of α-helical structure, this finding evidently showed the presence of α-helix and is consistent with the data from the conformation-dependent 13C NMR chemical shifts of corresponding 13C NMR signals. It turned out, therefore, that characterization of the α-helix structure based on REDOR experiment is straightforward and practical, if careful setting of experimental conditions is achieved to arrive at the meaningful data. It is also proved that no correction from the REDOR effect from the neighboring chains is necessary as far as α-helix structures are concerned, because fully and 30% labeled samples gave rise to identical REDOR results. Further, it is concluded that unusual displacement of 15N chemical shifts present in the sequential oligopeptide should be ascribed to a contribution other than any modified α-helical structures.