Side Chain Accessibility and Dynamics in the Molten Globule State of α-Lactalbumin: A19F-NMR Study

Abstract

The molten globule state of α-lactalbumin (α-LA) has been considered a prototype of partially folded proteins. Despite the importance of molten globules in understanding the mechanisms of protein folding and its relevance to some biological phenomena, site-specific information on the structure and dynamics of a molten globule is limited, largely because of the high conformational flexibility and heterogeneity. Here, we use selective isotope labeling and ¹⁹F NMR to investigate the solvent accessibility and side-chain dynamics of aromatic residues in the molten globule of α-LA. Comparison of these properties with those of the native and unfolded protein indicates that the α-LA molten globule is highly heterogeneous; each residue has its unique solvent accessibility and motional environment. Many aromatic residues normally buried in the interior of native α-LA remain significantly buried in the molten globule and the side-chain dynamics of these residues are highly restricted. Our results suggest that hydrophobic and van der Waals interactions mediated by the inaccessible surface area could be sufficient to account for all the stability of the α-LA molten globule, which is approximately 50% of the value for the native protein.