Quantitative analysis of helix-coil transition of block copolypeptide, Glu12-Ala12, by combined use of CD and NMR spectroscopy

To investigate helix-coil transition mechanisms, conformations of Glu(12)-Ala(12), EA, in aqueous solution have been studied in detail over the pH range from 2 to 8 and the temperature range from 20 to 60 degrees C using CD and NMR spectroscopy. The 750-MHz NMR spectra displayed excellent dispersion of the backbone amide proton signals, and permitted essentially complete sequence-specific resonance assignments. These assignments, together with short- and medium range nuclear Overhauser effect (NOE) constraints and coupling constants, enable us to analyze conformational characteristics of all the residues in the EA peptide individually. A combined use of CD and NMR techniques reveals that the EA peptide assumes a stable alpha-helix from Glu12 to Ala19 in 0. 1M NaCl solution at 20 degrees C above pH 7. The a-helix is getting longer as decreasing pH. Below pH 4, the peptide assumes the longest alpha-helix from Glu3 to Ala23. The important observation of the present study is that the helix- coil transition occurs stepwise, residue by residue, from both the Nand C-termini of the alpha-helix. No conformational equilibrium between the helical and random-coil states is detected for the residues in the central region of the a-helix. Quantitative analysis of temperature-induced helix-to-coil transitions at various pHs provides a pH-independent residual enthalpy change Delta H-r = 0.95 kcal res (-1). Similar values hove been reported for a 50-residue alanine-rich peptide (1.2 kcal res(-1)), poly-(L)-glutainate (1.1 kcal res(-1)), poly-(L)-lysine (1.1 kcal res(-) 1), and poly-(L)-alanine (0.86 kcal res(-1)). Those investigations, along with our present result, suggest that Delta H-r is mainly determined by the transformation of the backbone associated with the disruption of the intramolecular hydrogen bond. These results should increase our understanding of the helix-coil transition. (c) 2005 Wiley Periodicals, Inc.