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 <i>beta</i> and that are studied in water at 2 °C, pH 1−8. NMR analysis yields a structural characterization of the peptide Ac^βAspHelAla₈<i>beta</i>NH₂ and selected members of one ^βAspHelAla<i>_n</i><i>beta</i> 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<i>_n</i> hydrogen bonding signatures as α-helical without detectable 3₁₀ character. Relative NH→ND exchange rates yield site protection factors PF<i>_i</i> that define uniquely high fractional helicities FH for the peptide Ala<i>_n</i> regions. These Ala<i>_n</i> calibration series, studied in water and lacking helix-stabilizing tertiary structure, yield the first ¹³C NMR chemical shifts, ³<i>J</i>_HNH_α coupling constants, and CD ellipticities [θ_Molar]_λ_,<i>_n</i> characteristic of a fully helical alanine within an Ala<i>_n</i> context. CD data are used to assign parameters <i>X</i> and [θ]_λ_,_∞, required for rigorous calculation of FH values from CD ellipticities.