CONFORMATIONAL SPECIFICITY OF THE CHAPERONIN GROEL FOR THE COMPACT FOLDING INTERMEDIATES OF ALPHA-LACTALBUMIN

The chaperonin GroEL binds unfolded polypeptides, preventing aggregati on, and then mediates their folding in an ATP-dependent process. To un derstand the structural features in non-native polypeptides recognized by GroEL, we have used alpha-lactalbumin (alpha LA) as a model substr ate. alpha LA (14.2 kDa) is stabilized by four disulfide bonds and a b ound Ca2+ ion, offering the possibility of trapping partially folded d isulfide intermediates between the native and the fully unfolded state . The conformers of alpha LA with high affinity for GroEL are compact, containing up to three disulfide bonds, and have significant secondar y structure, but lack stable tertiary structure and expose hydrophobic surfaces. Complex formation requires almost the complete alpha LA seq uence and is strongly dependent on salts that stabilize hydrophobic in teractions. Unfolding of alpha LA to an extended state as well as the burial of hydrophobic surface upon formation of ordered tertiary struc ture prevent the binding to GroEL. Interestingly, GroEL interacts only with a specific subset of the many partially folded disulfide interme diates of alpha LA and thus may influence in vitro the kinetics of the folding pathways that lead to disulfide bonds with native combination s. We conclude that the chaperonin interacts with the hydrophobic surf aces exposed by proteins in a flexible compact intermediate or molten globule state.