title: A functional R domain from cystic fibrosis transmembrane conductance regulator is predominantly unstructured in solution.
abstract: Phosphorylation of the regulatory (R) domain initiates cystic fibrosis transmembrane conductance regulator (CFTR) Cl(-) channel activity. To discover how the function of this domain is determined by its structure, we produced an R domain protein (R8) that spanned residues 708-831 of CFTR. Phosphorylated, but not unphosphorylated, R8 stimulated activity of CFTR channels lacking this domain, indicating that R8 is functional. Unexpectedly, this functional R8 was predominantly random coil, as revealed by CD and limited proteolysis. The CD spectra of both phosphorylated and nonphosphorylated R8 were similar in aqueous buffer. The folding agent trimethylamine N-oxide induced only a small increase in the helical content of nonphosphorylated R8 and even less change in the helical content of phosphorylated R8. These data, indicating that the R domain is predominantly random coil, may explain the seemingly complex way in which phosphorylation regulates CFTR channel activity.
authors.affiliation: Howard Hughes Medical Institute, Department of Internal Medicine, University of Iowa College of Medicine, Iowa City, IA 52242, USA.
authors.forename: L S
authors.initials: LS
authors.lastname: Ostedgaard
authors.forename: O
authors.initials: O
authors.lastname: Baldursson
authors.forename: D W
authors.initials: DW
authors.lastname: Vermeer
authors.forename: M J
authors.initials: MJ
authors.lastname: Welsh
authors.forename: A D
authors.initials: AD
authors.lastname: Robertson
mesh_terms: D002942:Circular Dichroism; D003001:Cloning, Molecular; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D004926:Escherichia coli; D006801:Humans; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D010766:Phosphorylation; D017433:Protein Structure, Secondary; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017384:Sequence Deletion; D012996:Solutions
chemical_list: C505032:CFTR protein, human; D011994:Recombinant Proteins; D012996:Solutions; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D017868:Cyclic AMP-Dependent Protein Kinases