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

• Published in: Journal of the American Chemical Society 2005-02, Vol.127 (6), p.1690-1704
• Main Authors: Heitmann, Björn, Job, Gabriel E, Kennedy, Robert J, Walker, Sharon M, Kemp, Daniel S
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Calibration; Carbon Isotopes; Circular Dichroism - methods; Circular Dichroism - standards; Conformational dynamics in molecular biology; Fundamental and applied biological sciences. Psychology; Hydrogen-Ion Concentration; Models, Molecular; Molecular biophysics; Nuclear Magnetic Resonance, Biomolecular - methods; Peptides - chemistry; Protein Structure, Secondary; Protons; Solubility; Water - chemistry
• ISSN: 0002-7863; 1520-5126
• DOI: 10.1021/ja0457462

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 AcβAspHelAla8 betaNH2 and selected members of one βAspHelAla n beta 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, 3 J HNH α coupling constants, and CD ellipticities [θMolar]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.