Local interactions drive the formation of nonnative structure in the denatured state of human alpha-lactalbumin: A high resolution structural characterization of a peptide model in aqueous solution

There are a small number of peptides derived from proteins that have a prop ensity to adopt structure in aqueous solution which is similar to the struc ture they possess in the parent protein. There are far fewer examples of pr otein fragments which adopt stable nonnative structures in isolation. Under standing how nonnative interactions are involved in protein folding is cruc ial to our understanding of the topic. Here we show that a small, 11 amino acid peptide corresponding to residues 101-111 of the protein alpha-lactalb umin is remarkably structured in isolation in aqueous solution. The peptide has been characterized by H-1 NMR, and 170 ROE-derived constraints were us ed to calculate a structure. The calculations yielded a single, high-resolu tion structure for residues 101-107 that is nonnative in both the backbone and sidechain conformations. In the pH 6.5 crystal structure, residues 101- 105 are in an irregular turn-like conformation and residues 106-111 form an alpha-helix. In the pH 4.2 crystal structure, residues 101-105 form an alp ha-helix, and residues 106-111 form a loopike structure. Both of these stru ctures are significantly different from the conformation adopted by our pep tide. The structure in the peptide model is primarily the result of local s ide-chain interactions that force the backbone to adopt a nonnative 3(10)tu rn-like structure in residues 103-106. The structure in aqueous solution wa s compared to the structure in 30% trifluoroethanol (TFE), and clear differ ences were observed. In particular, one of the side-chain interactions, a h ydrophobic cluster involving residues 101-105, is different in the two solv ents and residues 107-111 are considerably more ordered in 30% TFE. The imp lications of the nonnative structure for the folding of alpha-lactalbumin i s discussed.