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J Biol Chem. 2018 Nov 16;293(46):17685-17704. doi: 10.1074/jbc.RA117.000819. Epub 2018 Jun 14.

Ligand binding to a remote site thermodynamically corrects the F508del mutation in the human cystic fibrosis transmembrane conductance regulator.

Author information

From the Departments of Biological Sciences and.
the Department of Biochemistry and Biophysics, School of Medicine, University of North Carolina, Chapel Hill, North Carolina 27599.
the Department of Chemistry, University of Alabama, Birmingham, Alabama 35294, and.
the Department of Chemistry, Bar-Ilan University, Ramat-Gan 5290002, Israel.
Chemistry, Columbia University, New York, New York 10027.
From the Departments of Biological Sciences and


Many disease-causing mutations impair protein stability. Here, we explore a thermodynamic strategy to correct the disease-causing F508del mutation in the human cystic fibrosis transmembrane conductance regulator (hCFTR). F508del destabilizes nucleotide-binding domain 1 (hNBD1) in hCFTR relative to an aggregation-prone intermediate. We developed a fluorescence self-quenching assay for compounds that prevent aggregation of hNBD1 by stabilizing its native conformation. Unexpectedly, we found that dTTP and nucleotide analogs with exocyclic methyl groups bind to hNBD1 more strongly than ATP and preserve electrophysiological function of full-length F508del-hCFTR channels at temperatures up to 37 °C. Furthermore, nucleotides that increase open-channel probability, which reflects stabilization of an interdomain interface to hNBD1, thermally protect full-length F508del-hCFTR even when they do not stabilize isolated hNBD1. Therefore, stabilization of hNBD1 itself or of one of its interdomain interfaces by a small molecule indirectly offsets the destabilizing effect of the F508del mutation on full-length hCFTR. These results indicate that high-affinity binding of a small molecule to a remote site can correct a disease-causing mutation. We propose that the strategies described here should be applicable to identifying small molecules to help manage other human diseases caused by mutations that destabilize native protein conformation.


F508del mutation; calorimetry; cystic fibrosis; cystic fibrosis transmembrane conductance regulator (CFTR); deoxythymidine; electrophysiology; fluorescence resonance energy transfer (FRET); high-throughput screening (HTS); isothermal titration calorimetry (ITC); protein aggregation; thermodynamics

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