Domain-interface dynamics of CFTR revealed by stabilizing nanobodies

Area of Science:

• Biochemistry
• Molecular Biology
• Structural Biology

Background:

• Cystic fibrosis (CF) is primarily caused by the F508del mutation in the cystic fibrosis transmembrane conductance regulator (CFTR) protein.
• This mutation destabilizes the NBD1 domain and disrupts its interaction with the transmembrane domain, leading to protein degradation.

Purpose of the Study:

• To develop nanobodies that target and stabilize the NBD1 domain of human CFTR.
• To elucidate the structural basis of NBD1 stabilization by nanobodies.

Main Methods:

• Development of nanobodies targeting human CFTR NBD1.
• Crystallography to determine the structure of NBD1-nanobody complexes.
• Biochemical assays to assess protein stabilization.

Main Results:

• Nanobodies successfully stabilized both isolated NBD1 and full-length CFTR protein.
• Crystal structures revealed specific epitopes and the molecular mechanisms of stabilization.
• A novel CFTR conformation with detached NBD1 was observed, differing from previously reported structures.

Conclusions:

• Nanobodies can effectively stabilize the CFTR NBD1 domain, offering a potential therapeutic avenue for CF.
• The identified CFTR conformation provides new insights into protein function and pathogenesis.
• Findings are relevant for understanding CFTR and other ABC transporter proteins.