Nucleation and stability of hydrogen-bond surrogate-based alpha-helices

Abstract

We have reported a new class of artificial alpha-helices in which a pre-organized alpha-turn nucleates the helical conformation [R. N. Chapman, G. Dimartino, and P. S. Arora, J Am. Chem. Soc., 2004, 126, 12252 and D. Wang, K. Chen, J. L. Kulp, III, and P. S. Arora, J. Am. Chem. Soc., 2006, 128, 9248]. This manuscript describes the effect of the core nucleation template on the overall helicity of the peptides and demonstrates that the macrocycle which most closely mimics the 13-membered hydrogen-bonded alpha-turn in canonical alpha-helices also affords the most stable artificial alpha-helix. We also investigate the stability of these synthetic helices through classical helix-coil parameters and find that the denaturation behavior of HBS alpha-helices agrees with the theoretical properties of a peptide with a well-defined and stable helix nucleus.

Fig. 1

Nucleation of short α-helices by replacement of an N-terminal i and i + 4 hydrogen bond with a carbon–carbon bond.

Fig. 2

(a) Determination of the optimum nucleation macrocycle in HBS helices. (b) Control peptides and HBS α-helices designed to determine the effect of the nucleation macrocycle on the overall helicity.

Fig. 3

Circular dichroism spectra of (a) 1 (black), 3a (red), 3b (green), 3c (pink), 3d (blue) and (b) 2 (black), 4a (red), 4b (green), 4c (pink), 4d (blue). The CD spectra were obtained in 10% TFE–PBS.

Fig. 4

NOESY correlation charts and cross-sections of NOESY spectra for 3a (a, c) and 3b (b, d). The alanine-3 residues in both artificial helices are N-alkylated. Filled rectangles indicate relative intensity of the NOE cross-peaks. Empty rectangles indicate NOE that could not be unambiguously assigned because of overlapping signals.

Fig. 5

Temperature-dependent chemical shifts (a, c) and hydrogen–deuterium exchange plots (b, d) for backbone amide protons in 3a and 3b.

Fig. 6

(a) Effect of temperature on the stability of HBS α-helices 3a and 3b. (b) Concentration dependence of mean residue ellipticity of HBS helices at 25 °C. The CD spectra were obtained in 10% TFE–PBS. (c) Normalized thermal denaturation curves for HBS helices. (d) Comparison of theoretical denaturation curves as a function of different nucleation constant, σ, values with the normalized experimental denaturation curve. The theoretical curves were obtained by simulating eqn (1) and (2).

Fig. 7

Comparison of HBS helices to a theoretical oligomer with one end always locked in the helix state (eq. 3). The experimental denaturation curve of HBS helices agrees well with the simulated curve from eq. 3.