Conformational properties of four peptides spanning the sequence of hen lysozyme

Four peptides encompassing the entire amino acid sequence of hen lysozyme were examined in aqueous solution and in 50% (v/v) 2,2,2-trifluoroethanol (TFE) by far-UV CD. Two peptides, 1-40 and 84-129, correspond to regions which are helical in the native protein, and together represent the alpha-domain. The beta-domain of the native enzyme was also synthesized as two peptides, one (41-60) containing the residues in the triple stranded antiparallel beta-sheet and the other (61-82) corresponding to a region lacking regular secondary structure. In water at pH 2.0 and 25 degrees C, the monomeric peptides 1-40, 41-60 and 61-82 appear to be predominantly unstructured. By contrast, the peptide 84-129 has considerable, presumably helical structure, corresponding to approximately 19%, or nine residues, on average, which can be unfolded by the addition of 8 M urea or 6 M guanidine hydrochloride. In 50% TFE the conformational properties of the four peptides are again distinct. Although little helical structure is induced in the peptides 41-60 and 61-82, and a native-like extent of helical structure is induced in the peptide 1-40, the peptide 84-29 converts almost entirely to helical structure in 50% TFE. The far-UV CD spectrum of a stoichiometric mixture of the four peptides in water resembles closely that of a denatured state of the intact protein formed by reductive methylation of its four disulphide bonds, but differs significantly from that of the native protein. The far-UV CD spectrum of the peptide mixture in TFE is indistinguishable from that of the intact protein in this solvent, both in the presence and in the absence of its four disulphide bonds. The conformational preferences of the peptides are not predicted using standard assessments of helical propensity or hydrophobicity, but correlate instead with the number of local contacts made in the native protein. On the basis of these results, we suggest that the region 84-129 could play an important role in determining the nature of the early folding events in the folding pathway of the intact polypeptide chain.