title: Computational studies reveal phosphorylation-dependent changes in the unstructured R domain of CFTR.
abstract: The cystic fibrosis transmembrane conductance regulator (CFTR) is a cAMP-dependent chloride channel that is mutated in cystic fibrosis, an inherited disease of high morbidity and mortality. The phosphorylation of its approximately 200 amino acid R domain by protein kinase A is obligatory for channel gating under normal conditions. The R domain contains more than ten PKA phosphorylation sites. No individual site is essential but phosphorylation of increasing numbers of sites enables progressively greater channel activity. In spite of numerous studies of the role of the R domain in CFTR regulation, its mechanism of action remains largely unknown. This is because neither its structure nor its interactions with other parts of CFTR have been completely elucidated. Studies have shown that the R domain lacks well-defined secondary structural elements and is an intrinsically disordered region of the channel protein. Here, we have analyzed the disorder pattern and employed computational methods to explore low-energy conformations of the R domain. The specific disorder and secondary structure patterns detected suggest the presence of molecular recognition elements (MoREs) that may mediate phosphorylation-regulated intra- and inter-domain interactions. Simulations were performed to generate an ensemble of accessible R domain conformations. Although the calculated structures may represent more compact conformers than occur in vivo, their secondary structure propensities are consistent with predictions and published experimental data. Equilibrium simulations of a mimic of a phosphorylated R domain showed that it exhibited an increased radius of gyration. In one possible interpretation of these findings, by changing its size, the globally unstructured R domain may act as an entropic spring to perturb the packing of membrane-spanning sequences that constitute the ion permeability pathway and thereby activate channel gating.
authors.affiliation: Department of Biochemistry and Biophysics, University of North Carolina-Chapel Hill, Chapel Hill, NC 27599, USA. hegedus@med.unc.edu
authors.forename: Tamás
authors.initials: T
authors.lastname: Hegedus
authors.forename: Adrian W R
authors.initials: AW
authors.lastname: Serohijos
authors.forename: Nikolay V
authors.initials: NV
authors.lastname: Dokholyan
authors.forename: Lihua
authors.initials: L
authors.lastname: He
authors.forename: John R
authors.initials: JR
authors.lastname: Riordan
mesh_terms: D000465:Algorithms; D000595:Amino Acid Sequence; D000818:Animals; D003198:Computer Simulation; D003550:Cystic Fibrosis / Q000235:genetics; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000737:chemistry* / Q000235:genetics; D006801:Humans; D015640:Ion Channel Gating; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009010:Monte Carlo Method; D010766:Phosphorylation; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary
chemical_list: D019005:Cystic Fibrosis Transmembrane Conductance Regulator