title: Specific stabilization of CFTR by phosphatidylserine.
abstract: The Cystic Fibrosis Transmembrane Conductance Regulator (CFTR, ABCC7) is a plasma membrane chloride ion channel in the ABC transporter superfamily. CFTR is a key target for cystic fibrosis drug development, and its structural elucidation would advance those efforts. However, the limited in vivo and in vitro stability of the protein, particularly its nucleotide binding domains, has made structural studies challenging. Here we demonstrate that phosphatidylserine uniquely stimulates and thermally stabilizes the ATP hydrolysis function of purified human CFTR. Among several lipids tested, the greatest stabilization was observed with brain phosphatidylserine, which shifted the Tm for ATPase activity from 22.7±0.8°C to 35.0±0.2°C in wild-type CFTR, and from 26.6±0.7°C to 42.1±0.2°C in a more stable mutant CFTR having deleted regulatory insertion and S492P/A534P/I539T mutations. When ATPase activity was measured at 37°C in the presence of brain phosphatidylserine, Vmax for wild-type CFTR was 240±60nmol/min/mg, a rate higher than previously reported and consistent with rates for other purified ABC transporters. The significant thermal stabilization of CFTR by phosphatidylserine may be advantageous in future structural and biophysical studies of CFTR.
authors.affiliation: Department of Cell Biology and Biochemistry, and Center for Membrane Protein Research, Texas Tech University Health Sciences Center, 3601 4th Street, Stop 6540, Lubbock, TX 79430, USA. Electronic address: ellen.hildebrandt@ttuhsc.edu.
authors.forename: Ellen
authors.initials: E
authors.lastname: Hildebrandt
authors.affiliation: Department of Chemistry, Bar-Ilan University, Ramat-Gan, 5290002, Israel. Electronic address: netalyk@gmail.com.
authors.forename: Netaly
authors.initials: N
authors.lastname: Khazanov
authors.affiliation: Department of Medicine, University of Alabama at Birmingham, 701 19th Street South, Birmingham, AL 35294-0007, USA; Birmingham Veterans Medical Center, Research Service, Birmingham, AL 35233, USA. Electronic address: kappesjc@uab.edu.
authors.forename: John C
authors.initials: JC
authors.lastname: Kappes
authors.affiliation: Department of Medicine, University of Alabama at Birmingham, 701 19th Street South, Birmingham, AL 35294-0007, USA. Electronic address: daiq@uab.edu.
authors.forename: Qun
authors.initials: Q
authors.lastname: Dai
authors.affiliation: Department of Chemistry, Bar-Ilan University, Ramat-Gan, 5290002, Israel. Electronic address: hanoch@biu.ac.il.
authors.forename: Hanoch
authors.initials: H
authors.lastname: Senderowitz
authors.affiliation: Department of Cell Biology and Biochemistry, and Center for Membrane Protein Research, Texas Tech University Health Sciences Center, 3601 4th Street, Stop 6540, Lubbock, TX 79430, USA. Electronic address: ina.urbatsch@ttuhsc.edu.
authors.forename: Ina L
authors.initials: IL
authors.lastname: Urbatsch
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D000251:Adenosine Triphosphatases; D000255:Adenosine Triphosphate / Q000378:metabolism; D001665:Binding Sites / Q000502:physiology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D009154:Mutation / Q000235:genetics; D009711:Nucleotides / Q000378:metabolism; D010718:Phosphatidylserines / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology
chemical_list: D018528:ATP-Binding Cassette Transporters; C505032:CFTR protein, human; D009711:Nucleotides; D010718:Phosphatidylserines; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases
keywords: ABC transporters; ATP hydrolysis; Blind docking; Cystic Fibrosis Transmembrane Conductance Regulator; Phosphatidylserine; Thermal stability