A defective flexible loop contributes to the processing and gating defects of the predominant cystic fibrosis-causing mutation

People with the genetic disease cystic fibrosis (CF) often carry a deletion mutation DF508 on the gene encoding the CF transmembrane conductance regulator (CFTR) Cl² channel. This mutation greatly reduces the CFTR maturation process and slows the channel opening rate. Here, we investigate whether residues near F508 contribute to these defects in DF508-CFTR. Most deletion mutations, but not alanine substitutions, of individual residues from positions 503 to 513 impaired CFTR maturation. Interestingly, only protein processing of DY512-CFTR, like that of DF508-CFTR, was greatly improved by low-temperature culture at 27°C or small-molecule corrector C18. The 2 mutant Cl² channels were equally slow to open, suggesting that they may share common structural flaws. Studies on the H3-H4 loop that links residues F508 and Y512 demonstrate that G509A/V510G mutations, moving G509 1 position backward in the loop, markedly enhanced DF508-CFTR maturation and opening rate while promoting protein stability and persistence of the H3 helix in DF508 nucleotide-binding domain 1. Moreover, V510A/S511A mutations noticeably increased DY512-CFTR maturation at 27°C and its opening rate. Thus, loop abnormalities may contribute to DF508- and DY512-CFTR defects. Importantly, correcting defects from G509 displacement in DF508-CFTR may offer a new avenue for drug discovery and CF treatments.