Nitrogen-15 chemical shift tensors and conformation of solid polypeptides containing 15N-labeled L-alanine residue by 15N NMR. 2. Secondary structure is reflected in .sigma.22

Abstract

Absbrct: A series of polypeptides [Ala*,X], containing I5N-labeled L-alanine (Ala*) and other amino acids (X; natural abundance of 15N) such as glycine, L-alanine, palanine, L-leucine, &benzyl L-aspartate, y-benzyl L-glutamate, y-methyl L-glutamate, L-valine, L-isoleucine, and sarcosine were synthesized by the a-amino acid N-carboxy anhydride (NCA) method. Conformations of these polypeptides in the solid state were characterized on the basis of conformation-dependent I3C chemical shifts in the I3C cross-polarization-magic-angle spinning (CP-MAS) NMR spectra and of the characteristic bands in the infrared (IR) and far-IR spectra. Further, isotropic 15N chemical shift (ab) and chemical shift tensors (ull, u2, and uaa) of the polypeptides were measured by the 15N CP-MAS and the 15N CP-static (powder pattern) methods, respectively. It was found that ai, is useful for the conformational study of homopolypeptides and copolypeptides with identical primary structures (amino acid sequences). In addition, it was demonstrated that the u22 value of the Ala* residue in copolypeptide is closely related to the main-chain conformations (such as the right-handed and left-handed a-helices, and the &sheet forms) rather than the amino acid sequence. Consequently, the uz2 value is very useful for conformational analysis of solid copolypeptides. Recently, high-resolution and solid-state 15N NMR has been increasingly applied to the investigation of polypeptides, proteins, and biopolymers.'-I2 This is primarily due to rapid progress in the development of both methodology and instrumentation. For example, sensitivity of the I5N resonance has been greatly improved by introduction of the cross-polarization magic-angle spinning (CP-MAS) methods. However, little attempt has yet been made to relate the 15N chemical shifts and chemical shift tensors to the structural parameters or primary, secondary, and higher ordered structures of synthetic polypeptides or natural proteins in the solid state. Furthermore, since a nitrogen atom possesses lone-pair electrons, it is of interest to examine the effects of the lone-pair electrons on the isotropic 15N chemical shift (P~) and the principal values of the I5N chemical shift tensors (ull, uZ2, and uj3 from the downfield side), which are a measure of the electronic structure. Thus, it is now expected that I5N NMR spectroscopy will contribute to and become increasingly important in the study on the structures and dynamics of synthetic polypeptides and natural proteins in the solid state. In our previous papers,I3J4 we have demonstrated that (1) the uho value determined by the ISN CP-MAS NMR method is very sensitive to the primary structure such as the variety of amino acid residue and the amino acid sequence, as well as the secondary structure (particular conformations) such as the a-helix and @-sheet forms of polypeptides in the solid state and (2) the ullr 92, and uj3 values determined by the CP-static (powder pattern) spectra for I5N-labeled copolypeptides are strongly influenced not only by the local conformations but also by the chemical nature of individual amino acid residues. In particular, it is noteworthy that one of the principal values, u22, may be useful for determination of the backbone conformation of polypeptides in the solid state. In this paper, therefore, we attempt to elucidate the origin of the individual components of the I5N chemical shift tensors of polypeptides in connection mainly with the primary structure such as the variety of amino acid residue and amino acid sequence and the secondary structure such as right-handed a-helix (a-helix), left-handed a-helix (aL-helixI5), and &sheet conformations. For this purpose, we have prepared a series of 15N-labeled polypeptides,