title: Characterizing diverse orthologues of the cystic fibrosis transmembrane conductance regulator protein for structural studies.
abstract: As an ion channel, the cystic fibrosis transmembrane conductance regulator (CFTR) protein occupies a unique niche within the ABC family. Orthologues of CFTR are extant throughout the animal kingdom from sharks to platypods to sheep, where the osmoregulatory function of the protein has been applied to differing lifestyles and diverse organ systems. In humans, loss-of-function mutations to CFTR cause the disease cystic fibrosis, which is a significant health burden in populations of white European descent. Orthologue screening has proved fruitful in the pursuit of high-resolution structural data for several membrane proteins, and we have applied some of the princples developed in previous studies to the expression and purification of CFTR. We have overexpressed this protein, along with evolutionarily diverse orthologues, in Saccharomyces cerevisiae and developed a purification to isolate it in quantities sufficient for structural and functional studies.
authors.affiliation: Faculty of Life Sciences, University of Manchester, Manchester M13 9PL, U.K.
authors.forename: Naomi L
authors.initials: NL
authors.lastname: Pollock
authors.affiliation: Faculty of Life Sciences, University of Manchester, Manchester M13 9PL, U.K.
authors.forename: Tracy L
authors.initials: TL
authors.lastname: Rimington
authors.affiliation: Faculty of Life Sciences, University of Manchester, Manchester M13 9PL, U.K. Robert.ford@manchester.ac.uk.
authors.forename: Robert C
authors.initials: RC
authors.lastname: Ford
mesh_terms: D000818:Animals; D016384:Consensus Sequence; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D003902:Detergents / Q000737:chemistry; D006031:Glycosylation; D006801:Humans; D008958:Models, Molecular; D009154:Mutation; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D010766:Phosphorylation; D010802:Phylogeny; D011487:Protein Conformation; D011499:Protein Processing, Post-Translational; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D012995:Solubility
chemical_list: C505032:CFTR protein, human; D003902:Detergents; D010446:Peptide Fragments; D011994:Recombinant Proteins; D019005:Cystic Fibrosis Transmembrane Conductance Regulator
keywords: CFTR; expression system; orthologues; purification