A pair of pyrene groups as a conformational probe for designed four-α-helix bundle polypeptides

Abstract

A 53-residue polypeptide designed to adopt a four-α-helix bundle structure has been synthesized. To probe the bundle structure, a pair of L-pyren-1-ylalanine (Pya) residues were introduced into two α-helix segments (the first and third α-helix segments or the second and third segments, respectively) in the 53-peptide. The peptides showed induced CD in the pyrene absorption region associated with the α-helical CD corresponding to the polypeptide main chain. The probe showed strong excimer emission in the fluorescence spectra, indicating the proximal orientation of the pyrene groups consistent with the formation of the bundle structure. Furthermore, both polypeptides showed similar CD spectra which were split at 350 nm (a positive peak at longer wavelength and a negative peak at shorter wavelength), suggesting that both pairs of pyrene rings on helical rods in parallel and antiparallel orientations are arranged in a right-handed sense in the bundle structure. Moreover, the pyrene excimer energy was efficiently transferred to dodecyl acridine orange, the efficiency of the transfer being due to the trapping of the dodecyl group within the polypeptide bundle.