STRUCTURAL HETEROGENEITY OF THE VARIOUS FORMS OF APOMYOGLOBIN - IMPLICATIONS FOR PROTEIN-FOLDING

Temperature-induced denaturation transitions of different structural f orms of apomyoglobin were studied monitoring intrinsic tryptophan fluo rescence. It was found that the tryptophans are effectively screened f rom solvent both in native and acid forms throughout most of the tempe rature range tested. Thus, the tryptophans' surroundings do not show a considerable change in structure where major protein conformational t ransitions have been found in apomyoglobin using other techniques. At high temperatures and under strong destabilizing conditions, the trypt ophans' fluorescence parameters show sigmoidal thermal denaturation. T hese results, combined with previous studies, show that the structure of this protein is heterogeneous, including native-like (tightly packe d) and molten globule-like substructures that exhibit conformation (de naturation) transitions under different conditions of pH and temperatu re (and denaturants). The results suggest that the folding of this pro tein proceeds via two ''nucleation'' events whereby native-like contac ts are formed. One of these events, which involves AGH ''core'' format ion, appears to occur very early in the folding process, even before s ignificant hydrophobic collapse in the rest of the protein molecule. F rom the current studies and other results, a rather detailed picture o f the folding of myoglobin is presented, on the level of specific stru ctures and their thermodynamical properties as well as formation kinet ics.