Nonintegral stoichiometry in CFTR gating revealed by a pore-lining mutation

Area of Science:

• Biochemistry
• Molecular Biology
• Ion Channel Physiology

Background:

• Cystic fibrosis transmembrane conductance regulator (CFTR) is an ATP-binding cassette (ABC) protein functioning as an ATP-gated chloride channel.
• CFTR gating is traditionally understood as strictly coupled to ATP hydrolysis, involving nucleotide-binding domain (NBD) dimerization and separation.

Purpose of the Study:

• To investigate the gating mechanism of CFTR and challenge the established coupling between ATP hydrolysis and channel gating.
• To characterize a novel mutant CFTR channel exhibiting distinct open states and altered gating kinetics.

Main Methods:

• Electrophysiological recordings of mutant CFTR channels in the presence of ATP.
• Analysis of gating transitions and ATP hydrolysis stoichiometry.

Main Results:

• A mutant CFTR channel was identified with two distinct open states (O1 and O2), with a preferred O1→O2 transition.
• This transition violates microscopic reversibility, indicating energy input from ATP hydrolysis.
• Multiple O1→O2 transitions within a single opening event suggest nonintegral ATP consumption per gating cycle.

Conclusions:

• CFTR gating does not strictly adhere to a one-to-one coupling with ATP hydrolysis, revealing a post-hydrolytic state.
• A six-state gating model is proposed, accommodating domain autonomy and allosteric regulation in CFTR function.