Water-Solubilized, Cap-Stabilized, Helical Polyalanines:  Calibration Standards for NMR and CD Analyses

NMR and CD studies are reported for two length series of solubilized, spaced, highly helical polyalanines that are N-capped by the optimal helix stabilizer ^βAsp-Hel and C-capped by β-aminoalanine beta and that are studied in water at 2 °C, pH 1−8. NMR analysis yields a structural characterization of the peptide Ac^βAspHelAla₈betaNH₂ and selected members of one ^βAspHelAla_nbeta series. At pH > 4.5 the ^βAspHel cap provides a preorganized triad of carboxylate anion and two amide residues that is complementary to the helical polyalanine N-terminus. The C-terminal β-aminoalanine assumes a helix-stabilizing conformation consistent with literature precedents. H(N)CO NMR experiments applied to capped, uniformly ¹³C- and ¹⁵N-labeled Ala₈ and Ala₁₂ peptides define Ala_n hydrogen bonding signatures as α-helical without detectable 3₁₀ character. Relative NH→ND exchange rates yield site protection factors PF_i that define uniquely high fractional helicities FH for the peptide Ala_n regions. These Ala_n calibration series, studied in water and lacking helix-stabilizing tertiary structure, yield the first ¹³C NMR chemical shifts, ³J_HNHα coupling constants, and CD ellipticities [θ_Molar]_λ,n characteristic of a fully helical alanine within an Ala_n context. CD data are used to assign parameters X and [θ]_λ,∞, required for rigorous calculation of FH values from CD ellipticities.