Identification of binding sites for ivacaftor on the cystic fibrosis transmembrane conductance regulator
Onofrio Laselva,
Zafar Qureshi,
Zhi-Wei Zeng,
Evgeniy V. Petrotchenko,
Mohabir Ramjeesingh,
C. Michael Hamilton,
Ling-Jun Huan,
Christoph H. Borchers,
Régis Pomès,
Robert Young,
Christine E. Bear
Affiliations
Onofrio Laselva
Programme in Molecular Medicine, Hospital for Sick Children, 686 Bay Street, Toronto, ON M5G 0A4, Canada; Department of Medical and Surgical Sciences, University of Foggia, Foggia, Italy
Zafar Qureshi
Department of Chemistry, Simon Fraser University, Burnaby, Canada
Zhi-Wei Zeng
Programme in Molecular Medicine, Hospital for Sick Children, 686 Bay Street, Toronto, ON M5G 0A4, Canada; Department of Biochemistry, University of Toronto, Toronto, Canada
Evgeniy V. Petrotchenko
Segal Cancer Proteomics Center, Lady Davis Institute, Jewish General Hospital, McGill University, Montreal, Canada; Center for Computational and Data-Intensive Science and Engineering, Skolkovo Institute of Science and Technology, Moscow 121205, Russia
Mohabir Ramjeesingh
Programme in Molecular Medicine, Hospital for Sick Children, 686 Bay Street, Toronto, ON M5G 0A4, Canada
C. Michael Hamilton
Department of Chemistry, Simon Fraser University, Burnaby, Canada
Ling-Jun Huan
Programme in Molecular Medicine, Hospital for Sick Children, 686 Bay Street, Toronto, ON M5G 0A4, Canada
Christoph H. Borchers
Segal Cancer Proteomics Center, Lady Davis Institute, Jewish General Hospital, McGill University, Montreal, Canada; Center for Computational and Data-Intensive Science and Engineering, Skolkovo Institute of Science and Technology, Moscow 121205, Russia; Gerald Bronfman Department of Oncology, Jewish General Hospital, McGill University, Montreal, Quebec H3T 1E2, Canada
Régis Pomès
Programme in Molecular Medicine, Hospital for Sick Children, 686 Bay Street, Toronto, ON M5G 0A4, Canada; Department of Biochemistry, University of Toronto, Toronto, Canada
Robert Young
Department of Chemistry, Simon Fraser University, Burnaby, Canada
Christine E. Bear
Programme in Molecular Medicine, Hospital for Sick Children, 686 Bay Street, Toronto, ON M5G 0A4, Canada; Department of Physiology, University of Toronto, Toronto, Canada; Department of Biochemistry, University of Toronto, Toronto, Canada; Corresponding author
Summary: Ivacaftor (VX-770) was the first cystic fibrosis transmembrane conductance regulator (CFTR) modulatory drug approved for the treatment of patients with cystic fibrosis. Electron cryomicroscopy (cryo-EM) studies of detergent-solubilized CFTR indicated that VX-770 bound to a site at the interface between solvent and a hinge region in the CFTR protein conferred by transmembrane (tm) helices: tm4, tm5, and tm8. We re-evaluated VX-770 binding to CFTR in biological membranes using photoactivatable VX-770 probes. One such probe covalently labeled CFTR at two sites as determined following trypsin digestion and analysis by tandem-mass spectrometry. One labeled peptide resides in the cytosolic loop 4 of CFTR and the other is located in tm8, proximal to the site identified by cryo-EM. Complementary data from functional and molecular dynamic simulation studies support a model, where VX-770 mediates potentiation via multiple sites in the CFTR protein.
