Contribution of individual residues to formation of the native-like tertiary topology in the alpha-lactalbumin molten globule.
Molten globules are partially folded forms of proteins that have native-like secondary structure and a compact geometry, but often without rigid, specific side-chain packing. Recently, the molten globule of alpha-lactalbumin (alpha-LA) has been shown to adopt a native-like tertiary topology, mainly localized in the alpha-helical domain. This native-like topology is reflected by the high effective concentration (Ceff) for formation of the 28-111 disulfide bond, which is approximately 10 to 40 times higher than the Ceff for formation of any non-native disulfide bond in the alpha-helical domain. In order to understand the mechanism for formation of the native-like tertiary topology, we substituted alanine for each of the 23 buried residues in the alpha-helical domain of alpha-LA and determined the effect of these substitutions on the Ceff for formation of the 28-111 disulfide bond. Our results indicate that a subset of hydrophobic residues is most important for formation of the native-like topology. These residues form a densely packed core in the three-dimensional structure of alpha-LA. In contrast, the less important residues consist of both hydrophobic and hydrophilic amino acids located at peripheral positions. These results suggest that a relatively small number of hydrophobic residues may be sufficient for specifying the overall structure of a protein during early stages of protein folding.