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13C and 15N Chemical Shielding Tensors in
Crystallographically Characterized Peptides We have used natural abundance 13C and 15N 2D-PASS experiments to determine chemical shielding tensors for all C and N sites in a systematic set of tripeptides. Peptides studied have known x-ray structures allowing direct structural correlation with isotropic shifts, shielding anisotropies and asymmetries. Tripeptides examined include a series with central Gly residues containing torsion angles corresponding to the standard protein secondary structures (a-helices and b-strands). As seen in solution NMR of proteins, the C isotropic shift for alpha helices is approximately 1 ppm downfield of that for beta strands. Similar correlations can be made for the anisotropy (szz - sxx) and the asymmetry (syy - sxx), however, the effects observed are 5- to 10-fold larger. This suggests that measurements of shielding tensors in protein C carbons is a facile tool for secondary structure determination. Variations in the isotropic shifts, anisotropies and asymmetries for carbonyl tensors are similar in size to that seen for C, however, there is no obvious correlation with secondary structure. Aromatic carbons in Phe and Tyr residues show substantial variation in isotropic shifts but small variations in anisotropy and asymmetry. Moreover, the 6 nonequivalent methyl groups in the asymmetric unit of crystalline Ala-Ala-Ala containing two peptides of slightly different conformation are resolved in the solid state MAS experiment. Typically, polymorphs of the same peptide show distinct spectra. These observations suggest that MAS NMR may be a useful technique for understanding hydrophobic forces in proteins and polypeptides. 15N MAS experiments of the peptides studied to date reveal the following useful feature: the amide tensors have small asymmetries and anisotropies in the range of 140 ppm to 160 ppm (Gly, Ala, Leu, Phe). The triple resonance MAS probe constructed for these 2-D studies makes use of relatively small samples (25-50 mg) and will be described. |