THE MOLECULAR CHAPERONE HSC70 ASSISTS THE IN-VITRO FOLDING OF THE N-TERMINAL NUCLEOTIDE-BINDING DOMAIN OF THE CYSTIC-FIBROSIS TRANSMEMBRANECONDUCTANCE REGULATOR

The most common disease-causing mutation in the cystic fibrosis transm embrane conductance regulator is a single amino acid deletion (Delta F 508) in the N-terminal cytosolic nucleotide-binding domain (NBD1), Thi s mutation has previously been shown to be a temperature-sensitive fol ding mutation that alters the folding pathway but not the native state stability of the isolated domain (Qu, B.-H., and Thomas, P. J. (1996) J. Biol. Chem, 271, 7261-7264), Here we provide evidence that the mol ecular chaperone Hsc70 productively interacts with NBD1 to increase th e folding yield of the domain and inhibit off-pathway associations lea ding to the formation of high molecular weight aggregates, Furthermore , we have sublocalized a region within NBD1 where Hsc70 binds, Notably , inhibition of NBD1 aggregation is not dependent upon the presence of Hsc70 in the early stages of folding, indicating that the chaperone m ay act on a folding intermediate. In the presence of K+ and Mg2+-ATP, conditions where Hsp70 binds substrate rapidly and can release it, Hsc 70 is less effective at inhibiting NBD1 aggregation, Thus, the rate of release of unfolded substrate is an important factor in preventing ag gregation and promoting folding of the domain, These results demonstra te that Hsc70 promotes the otherwise inefficient folding of Delta F-NB D1 and provide insight into the mechanisms by which molecular chaperon es assist proteins in folding.