title: Crystallogenesis of Membrane Proteins Mediated by Polymer-Bounded Lipid Nanodiscs.
abstract: For some membrane proteins, detergent-mediated solubilization compromises protein stability and functionality, often impairing biophysical and structural analyses. Hence, membrane-protein structure determination is a continuing bottleneck in the field of protein crystallography. Here, as an alternative to approaches mediated by conventional detergents, we report the crystallogenesis of a recombinantly produced membrane protein that never left a lipid bilayer environment. We used styrene-maleic acid (SMA) copolymers to solubilize lipid-embedded proteins into SMA nanodiscs, purified these discs by affinity and size-exclusion chromatography, and transferred proteins into the lipidic cubic phase (LCP) for in meso crystallization. The 2.0-Å structure of an α-helical seven-transmembrane microbial rhodopsin thus obtained is of high quality and virtually identical to the 2.2-Å structure obtained from traditional detergent-based purification and subsequent LCP crystallization.
authors.affiliation: Structural Neurobiology, Department of Biochemistry, University of Toronto, 1 King's College Circle, Toronto, ON M5S 1A8, Canada. Electronic address: jana.broecker@utoronto.ca.
authors.forename: Jana
authors.initials: J
authors.lastname: Broecker
authors.affiliation: Structural Neurobiology, Department of Biochemistry, University of Toronto, 1 King's College Circle, Toronto, ON M5S 1A8, Canada.
authors.forename: Bryan T
authors.initials: BT
authors.lastname: Eger
authors.affiliation: Structural Neurobiology, Department of Biochemistry, University of Toronto, 1 King's College Circle, Toronto, ON M5S 1A8, Canada; Department of Molecular Genetics, University of Toronto, 1 King's College Circle, Toronto, ON M5S 1A8, Canada. Electronic address: oliver.ernst@utoronto.ca.
authors.forename: Oliver P
authors.initials: OP
authors.lastname: Ernst
mesh_terms: D001436:Bacteriorhodopsins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray / Q000379:methods*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006216:Halobacteriaceae / Q000737:chemistry*; D008298:Maleates / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D049329:Nanostructures / Q000737:chemistry*; D011137:Polystyrenes / Q000737:chemistry*; D000072756:Protein Conformation, alpha-Helical; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012995:Solubility
chemical_list: D008298:Maleates; D008565:Membrane Proteins; D011137:Polystyrenes; D011994:Recombinant Proteins; C024074:styrene-maleic acid polymer; D001436:Bacteriorhodopsins
keywords: SMA; SMALP; crystallization; crystallogenesis; lipid nanodisc; lipidic cubic phase; membrane protein; polymer; protein structure; styrene-maleic acid copolymer