title: Expression Screening of Integral Membrane Proteins by Fusion to Fluorescent Reporters.
abstract: The production of recombinant integral membrane proteins for structural and functional studies remains technically challenging due to their relatively low levels of expression. To address this problem, screening strategies have been developed to identify the optimal membrane sequence and expression host for protein production. A common approach is to genetically fuse the membrane protein to a fluorescent reporter, typically Green Fluorescent Protein (GFP) enabling expression levels, localization and detergent solubilisation to be assessed. Initially developed for screening the heterologous expression of bacterial membrane proteins in Escherichia coli, the method has been extended to eukaryotic hosts, including insect and mammalian cells. Overall, GFP-based expression screening has made a major impact on the number of membrane protein structures that have been determined in the last few years.
authors.affiliation: OPPF-UK, The Research Complex at Harwell, Rutherford Appleton Laboratory Harwell, Oxford, UK.|Division of Structural Biology, Henry Wellcome Building for Genomic Medicine, University of Oxford, Roosevelt Drive, Oxford, UK.
authors.forename: Louise E
authors.initials: LE
authors.lastname: Bird
authors.affiliation: OPPF-UK, The Research Complex at Harwell, Rutherford Appleton Laboratory Harwell, Oxford, UK.|Division of Structural Biology, Henry Wellcome Building for Genomic Medicine, University of Oxford, Roosevelt Drive, Oxford, UK.
authors.forename: Joanne E
authors.initials: JE
authors.lastname: Nettleship
authors.affiliation: OPPF-UK, The Research Complex at Harwell, Rutherford Appleton Laboratory Harwell, Oxford, UK.|Division of Structural Biology, Henry Wellcome Building for Genomic Medicine, University of Oxford, Roosevelt Drive, Oxford, UK.
authors.forename: Valtteri
authors.initials: V
authors.lastname: Järvinen
authors.affiliation: OPPF-UK, The Research Complex at Harwell, Rutherford Appleton Laboratory Harwell, Oxford, UK.|Division of Structural Biology, Henry Wellcome Building for Genomic Medicine, University of Oxford, Roosevelt Drive, Oxford, UK.
authors.forename: Heather
authors.initials: H
authors.lastname: Rada
authors.affiliation: OPPF-UK, The Research Complex at Harwell, Rutherford Appleton Laboratory Harwell, Oxford, UK.|Division of Structural Biology, Henry Wellcome Building for Genomic Medicine, University of Oxford, Roosevelt Drive, Oxford, UK.
authors.forename: Anil
authors.initials: A
authors.lastname: Verma
authors.affiliation: OPPF-UK, The Research Complex at Harwell, Rutherford Appleton Laboratory Harwell, Oxford, UK. ray@strubi.ox.ac.uk.|Division of Structural Biology, Henry Wellcome Building for Genomic Medicine, University of Oxford, Roosevelt Drive, Oxford, UK. ray@strubi.ox.ac.uk.
authors.forename: Raymond J
authors.initials: RJ
authors.lastname: Owens
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D004926:Escherichia coli / Q000378:metabolism; D005057:Eukaryotic Cells / Q000378:metabolism; D015870:Gene Expression; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000032:analysis; D057809:HEK293 Cells / Q000378:metabolism; D006801:Humans; D007313:Insecta / Q000166:cytology; D008164:Luminescent Proteins / Q000032:analysis* / Q000235:genetics; D008565:Membrane Proteins / Q000032:analysis* / Q000096:biosynthesis / Q000235:genetics; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000096:biosynthesis / Q000235:genetics; D015003:Yeasts / Q000378:metabolism
chemical_list: D008164:Luminescent Proteins; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins
keywords: Escherichia coli; Green fluorescent protein; HEK 293 cells; Insect cells; Integral membrane protein; Pichia pastoris; Saccharomyces cerevisiae