Determination of chemical shielding tensor of an indole carbon and application to tryptophan orientation of a membrane peptide

Using tryptophan ¹³C-enriched at the C₄ (Cε₃) of the indole, the orientation of the Cε₃ chemical shift tensor relative to the Cε₃-H dipolar axis was determined from the ¹³C chemical shift/¹³C-¹H dipolar 2D NMR powder pattern. The principal values obtained were 208, 137 and 15 ppm with σ₃₃ perpendicular to the indole plane, and σ₁₁ (least shielded direction) 5° off the Cε₃-H bond toward C_ξ3. The side off the Cε₃-H bond was determined by comparing the reduced chemical shift anisotropies obtained by solid-state NMR and from molecular dynamics calculations of [4-¹³C] tryptophans in gramicidin A aligned in phospholipid membranes.