Timing of CFTR pore opening and structure of its transition state.
In CFTR, the chloride ion channel mutated in cystic fibrosis (CF) patients, pore opening is coupled to ATP-binding-induced dimerization of two cytosolic nucleotide binding domains (NBDs) and closure to dimer disruption following ATP hydrolysis. CFTR opening rate, unusually slow because of its high-energy transition state, is further slowed by CF mutation ΔF508. Here, we exploit equilibrium gating of hydrolysis-deficient CFTR mutant D1370N and apply rate-equilibrium free-energy relationship analysis to estimate relative timing of opening movements in distinct protein regions. We find clear directionality of motion along the longitudinal protein axis and identify an opening transition-state structure with the NBD dimer formed but the pore still closed. Thus, strain at the NBD/pore-domain interface, the ΔF508 mutation locus, underlies the energetic barrier for opening. Our findings suggest a therapeutic opportunity to stabilize this transition-state structure pharmacologically in ΔF508-CFTR to correct its opening defect, an essential step toward restoring CFTR function.
Department of Medical Biochemistry, Semmelweis University, Tűzoltó u. 37-47, Budapest 1094, Hungary; MTA-SE Ion Channel Research Group, Semmelweis University, Tűzoltó u. 37-47, Budapest 1094, Hungary.
Ben
B
Sorum
MTA-SE Ion Channel Research Group, Semmelweis University, Tűzoltó u. 37-47, Budapest 1094, Hungary.
Dávid
D
Czégé
Department of Medical Biochemistry, Semmelweis University, Tűzoltó u. 37-47, Budapest 1094, Hungary; MTA-SE Ion Channel Research Group, Semmelweis University, Tűzoltó u. 37-47, Budapest 1094, Hungary. Electronic address: csanady.laszlo@med.semmelweis-univ.hu.
László
L
Csanády
D000255:Adenosine Triphosphate / Q000378:metabolism; D020134:Catalytic Domain; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006801:Humans; D009154:Mutation; D017434:Protein Structure, Tertiary; D013816:Thermodynamics
C505032:CFTR protein, human; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000255:Adenosine Triphosphate