Structural basis for CFTR inhibition by CFTR_inh-172

Area of Science:

• Molecular Biology
• Structural Biology
• Biophysics

Background:

• The cystic fibrosis transmembrane conductance regulator (CFTR) is crucial for electrolyte and fluid balance.
• CFTR activation treats cystic fibrosis, while its inhibition is a therapeutic target for secretory diarrhea and polycystic kidney disease.
• The precise mechanism of action for many CFTR inhibitors remains unclear.

Purpose of the Study:

• To determine the structure of CFTR in complex with the inhibitor CFTR_inh-172.
• To elucidate the molecular mechanism by which CFTR_inh-172 inhibits CFTR channel activity.
• To provide a structural basis for the function of a widely used CFTR inhibitor.

Main Methods:

• Cryogenic electron microscopy (cryo-EM) to determine the high-resolution structure of CFTR-CFTR_inh-172 complex.
• Single-molecule fluorescence resonance energy transfer (smFRET) experiments to assess channel gating and nucleotide-binding domain dimerization.

Main Results:

• The 2.7 Å cryo-EM structure revealed CFTR_inh-172 binding within the CFTR pore near transmembrane helix 8.
• CFTR_inh-172 binding stabilizes a conformation with a collapsed chloride selectivity filter, blocking the pore from the extracellular side.
• smFRET experiments confirmed that CFTR_inh-172 inhibits channel gating without affecting nucleotide-binding domain dimerization.

Conclusions:

• The study provides a detailed molecular understanding of CFTR_inh-172 binding and inhibition.
• The findings reconcile previous biophysical data and explain the mechanism of this CFTR inhibitor.
• This structural insight is vital for the rational design of novel CFTR-targeting therapeutics.