CONFORMATIONAL PROPERTIES OF 4 PEPTIDES SPANNING THE SEQUENCE OF HEN LYSOZYME

Four peptides encompassing the entire amino acid sequence of hen lysoz yme were examined in aqueous solution and in 50% (v/v) 2,2,2-trifluoro ethanol (TFE) by far-UV CD. Two peptides, 1-40 and 84-129, correspond to regions which are helical in the native protein, and together repre sent the alpha-domain. The beta-domain of the native enzyme was also s ynthesized as two peptides, one (41-60) containing the residues in the triple stranded antiparallel beta-sheet and the other (61-82) corresp onding 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 ap pear to be predominantly unstructured. By contrast, the peptide 84-129 has considerable, presumably helical structure, corresponding to simi lar to 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 c onformational properties of the four peptides are again distinct. Alth ough 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 pe ptide 1-40, the peptide 84-129 converts almost entirely to helical str ucture in 50% TFE. The far-UV CD spectrum of a stoichiometric mixture of the four peptides in water resembles closely that of a denatured st ate of the intact protein formed by reductive methylation of its four disulphide bonds, but differs significantly from that of the native pr otein. The far-UV CD spectrum of the peptide mixture in TFE is indisti nguishable from that of the intact protein in this solvent, both in th e presence and in the absence of its four disulphide bonds. The confor mational preferences of the peptides are not predicted using standard assessments of helical propensity or hydrophobicity, but correlate ins tead 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 pl ay an important role in determining the nature of the early folding ev ents in the folding pathway of the intact polypeptide chain. (C) 1995 Academic Press Limited