Abstract
The binding constants between disulfide-intact or various disulfide-reduced bovine α-lactalbumins and an Escherichia coli chaperonin, GroEL, were determined by using the equilibrium dialysis method. The disulfide-intact and one-disulfide (Cys6-Cys120)-reduced a-lactalbumins were shown not to bind with GroEL both in the presence and absence of Call. The two-disulfide (Cys6-Cys120 and Cys28-Cys111)-reduced α-lactalbumin, which has the native-like tertiary structure in its β-domain region and an unfolded α-domain in the presence of Call, showed considerable binding with GroEL. The binding free energy of the two-disulfide-reduced a-lactalbumin in the presence of Call is close to that of the molten globule state of disulfide-intact α-lactalbumin. This result suggests that GroEL binds to the unfolded α-domain of α-lactalbumin regardless of the conformation of the β-domain. The fully disulfide-reduced and two-disulfide-reduced α-lactalbumins were found to bind more strongly with GroEL in the absence of Ca2+ than the two-disulfidereduced α-lactalbumin in the presence of Ca2+, thus indicating that the unfolding of the β-domain of a-lactalbumin leads to stronger interaction with GroEL.
