Allosteric Coupling Between the CFTR Chloride Channel's NBD1 Heterodimer Interface and Intracellular Domains

Abstract

Cystic fibrosis is caused by mutations in the chloride channel CFTR, leading to loss of function and changes in the ion and fluid flow across epithelial surfaces. Like ABC transporters, CFTR contains two membrane spanning domains (MSDs) and two cytoplasmic nucleotide binding domains (NBDs). The formation of an NBD1/NBD2 dimer drives channel opening. The coupling helices at the base of the intracellular domains (ICLs) couple the NBDs to the MSDs of the channel. How are changes on the heterodimer interface transmitted across NBD1 to ICLs? The sensitivity of NMR spectroscopy reveals how the ICL4 binding site of NBD1 is allosterically linked to its heterodimer interface. During titrations, an ICL4 “coupling helix” peptide bound near the alpha-subdomain of NBD1, leading to destabilization and release of the C-terminal NBD1 helices 8 and 9 (H8/H9) from the heterodimer interface via an allosteric mechanism. Therefore, perturbations in one region should cause a reciprocal change in the other region. DelF508, a CF-causing mutation in the alpha-subdomain, reduces the effects of ICL4 binding on H8/H9. In contrast, DelF508-suppressor mutations, F494N and V510D, increase these effects. Helix 8 mutation, Q637R (also a DelF508-suppressor), increases the binding effects in the ICL4 binding site. Q637R also alters the dynamics in this region, suggesting that the internal motions of NBD1 are involved in transmitting changes across this plastic domain. The destabilization and release of H8/H9 from the heterodimer interface is strikingly similar to that of the regulatory extension (RE) and R region, which follow helix 9 and become more disordered and less bound to NBD1 upon phosphorylation. The RE and R region regulate NBD dimerization and, ultimately, channel opening and closing. The allosteric pathway provides insight into how dimerization may be communicated to the rest of CFTR.