DETECTION AND CHARACTERIZATION OF INTERMEDIATE ON FOLDING PATHWAY OF HUMAN ALPHA-LACTALBUMIN

To discuss the relation between the folding mechanism and the chemical structure of proteins, the reversible unfolding reactions of human α-lactalbumin by acidification and by guanidine hydrochloride at 25 °C are studied by means of circular dichroism, difference spectra and pH-jump measurements and are compared with those for bovine α-lactalbumin. As shown previously for bovine α-lactalbumin, the folding process at neutral pH is not explained by a simple two-state mechanism but involves an intermediate form that has the same amount of helical structures as the native form. The transition between the intermediate and the fully denatured states is too rapid to be measured and corresponds to the helix-coil transition of the backbone. One of the differences of human α-lactalbumin from the bovine protein is the remarkable stability of the intermediate at neutral pH, which can be explained by differences in the primary chemical structure. Another difference is the existence at acid pH of an additional helical form, which is more helical than the native form. The transition from this to the intermediate or to the fully denatured one also is shown to resemble the helix-coil transition. The following folding scheme of human α-lactalbumin is proposed: FORMULA OMMITTED Here N is the native form, and the intermediate is a macroscopic state distributed around the state A0 at neutral pH, while the distribution in the acid and fully denatured states shifts toward Am and A_n respectively. © 1978, American Chemical Society. All rights reserved.