15N chemical-shift tensor of the imide nitrogen in the alanyl-prolyl peptide bond

Abstract

In order to extract the information about molecular structure and dynamics that is inherent in the chemical-shift interaction, it is necessary to know the magnitudes and orientations of the principal elements of the chemical-shift tensor for each site of interest. While the 13C and “N chemical-shift tensors for most of the chemical groups in proteins have been described, the sites in the imino acid proline have not yet been characterized. This note describes the chemical-shift tensor for the “N imide nitrogen in the alanyl-prolyl peptide bond based on the analysis of 15N powder patterns resulting from the chemical shift and heteronuclear dipole-dipole coupling in the 13C,‘5N-labeled dipeptide Ala-Pro. The magnitudes and orientations of the principal elements of the chemical-shift tensor are generally determined from rotation studies of single crystals (I, 2). Amino acids and small synthetic peptides are useful as model systems for larger and less tractable biologically important peptides and proteins. The 13C and “N chemical-shift tensors of the amide peptide bond have been determined in single crystal studies of the dipeptide glycyl-glycine (3, 4). The magnitudes of the principal values of the chemical-shift tensor can be readily determined from the frequencies of the discontinuities of a powder pattern obtained from a polycrystalline sample, as long as the site of interest gives a well-defined and resolved powder pattern. However, the orientations of the principal elements in the molecular frame cannot be determined from the powder pattern of the chemical-shift interaction alone. If there is a strong dipole-dipole coupling to a bonded atom, then it is possible to establish the orientation of the principal axis system relative to the bond axis in the molecular frame from the perturbation of the chemical-shift powder pattern by the dipolar splittings associated with each orientation in the sample. This was first demonstrated for i3C coupled to i5N by Kaplan et al. (5) and has been subsequently applied to 13C coupled to ‘H (6), 15N coupled to ‘H (7) and 13C coupled to 13c (8). Proteins of bacterial origin can be readily labeled with 15N in backbone and side chain sites for solid-state NMR studies (9). Proline is present in most proteins and it forms an imide linkage in the protein backbone. The i5N imide chemical-shift tensor