Structural characterization of the molten globule and native states of apomyoglobin by solution X-ray scattering

Compactness and shape are two of the critical properties that describe the degree of protein folding. Solution X-ray scattering is an effective technique for measuring these properties quantitatively. Structural characteristics of various conformational states of horse myoglobin were studied in terms of size and shape by solution X-ray scattering. The radius of gyration for native holomyoglobin was 17.5 A, while that of the apomyoglobin native state was 19.7 A. Corresponding to the increase in the radius of gyration, the largest dimension of the molecule also increased from 47.5 A to 62.5 A. Both states are globular in shape. The scattering profiles in the high angle region suggest that the apomyoglobin native state has a distinct tertiary structure, and that packing of alpha-helices in the apomyoglobin native state would be looser than that of holomyoglobin. These observations indicate that the native state of apomyoglobin is expanded from that of holomyoglobin, and that the conformations of the two are not identical. The radii of gyration for the acid-unfolded state and the denaturant-unfolded state were 30 A and 35 A, respectively. Both unfolded states have chain-like conformations without any tertiary structures. The radius of gyration and the largest dimension of the molten globule stabilized by trichloroacetate were 23.1 A and 72.5 A, respectively. The molten globule is expanded from the native state although it is globular, and is much more compact than the unfolded state. The bimodal distance distribution function and scattering profile at high-angle region suggest that the structure of the apomyoglobin molten globule contains a core comprising a cluster of multiple alpha-helices and flaring tail(s), which would be a common structural property of the compact denatured state appearing during the folding process. The compactness of each conformational state is highly correlated with the extent of formation of the alpha-helix.