ALTERATION OF THE CYSTIC-FIBROSIS TRANSMEMBRANE CONDUCTANCE REGULATORFOLDING PATHWAY - EFFECTS OF THE DELTA-F508 MUTATION ON THE THERMODYNAMIC STABILITY AND FOLDING YIELD OF NBD1

The cellular phenotype of the most common cystic fibrosis-causing muta tion, deletion of phenylalanine 508 (Delta F508) in the amino-terminal nucleotide binding domain (NBD1) of the cystic fibrosis transmembrane conductance regulator (CFTR), is the inability of the mutant protein to fold and transit to the apical membrane of affected epithelial cell s. Expressed NBD1s were purified and folded in vitro into soluble mono mers capable of binding nucleotide. Here we report that the Delta F508 mutation has little effect on the thermodynamic stability of the fold ed NBD1. The Delta G(D,GdnHCl)(0) is 15.5 kJ/mol for the wild type NBD 1 and 14.4 kJ/mol for NBD1 Delta F. In contrast, the mutation signific antly reduces the folding yield at a variety of temperatures, indicati ng that Phe-508 makes crucial contacts during the folding process, but plays little role in stabilization of the native state. Under conditi ons that approximate the efficiency of maturation in vivo, the rate of f-pathway is significantly increased by the disease causing mutation. These results establish a molecular mechanism for most cases of cystic fibrosis and provide insight into the complex processes by which prim ary sequence encodes the three-dimensional structure.