title: Controlled Co-reconstitution of Multiple Membrane Proteins in Lipid Bilayer Nanodiscs Using DNA as a Scaffold.
abstract: Nanodiscs constitute a tool for the solubilization of membrane proteins in a lipid bilayer, thus offering a near-native membrane environment. Many membrane proteins interact with other membrane proteins; however, the co-reconstitution of multiple membrane proteins in a single nanodisc is a random process that is adversely affected by several factors, including protein aggregation. Here, we present an approach for the controlled co-reconstitution of multiple membrane proteins in a single nanodisc. The temporary attachment of designated oligonucleotides to individual membrane proteins enables the formation of stable, detergent-solubilized membrane protein complexes by base-pairing of complementary oligonucleotide sequences, thus facilitating the insertion of the membrane protein complex into nanodiscs with defined stoichiometry and composition. As a proof of principle, nanodiscs containing a heterodimeric and heterotrimeric membrane protein complex were reconstituted using a fluorescently labeled voltage-gated anion channel (VDAC) as a model system.
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School , Boston, Massachusetts 02115, United States.
authors.forename: Thomas
authors.initials: T
authors.lastname: Raschle
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School , Boston, Massachusetts 02115, United States.|Department of Cancer Biology, Dana-Farber Cancer Institute , Boston, Massachusetts 02115, United States.|Wyss Institute for Biologically Inspired Engineering, Harvard University , Boston, Massachusetts 02115, United States.
authors.forename: Chenxiang
authors.initials: C
authors.lastname: Lin
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School , Boston, Massachusetts 02115, United States.|Department of Cancer Biology, Dana-Farber Cancer Institute , Boston, Massachusetts 02115, United States.|Wyss Institute for Biologically Inspired Engineering, Harvard University , Boston, Massachusetts 02115, United States.
authors.forename: Ralf
authors.initials: R
authors.lastname: Jungmann
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School , Boston, Massachusetts 02115, United States.|Department of Cancer Biology, Dana-Farber Cancer Institute , Boston, Massachusetts 02115, United States.|Wyss Institute for Biologically Inspired Engineering, Harvard University , Boston, Massachusetts 02115, United States.
authors.forename: William M
authors.initials: WM
authors.lastname: Shih
authors.affiliation: Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School , Boston, Massachusetts 02115, United States.
authors.forename: Gerhard
authors.initials: G
authors.lastname: Wagner
mesh_terms: D004247:DNA / Q000737:chemistry*; D005456:Fluorescent Dyes / Q000737:chemistry; D007473:Ion Channels / Q000737:chemistry; D008051:Lipid Bilayers / Q000737:chemistry*; D008565:Membrane Proteins / Q000737:chemistry*; D008856:Microscopy, Fluorescence; D008954:Models, Biological; D049329:Nanostructures / Q000737:chemistry*; D015530:Nuclear Matrix / Q000737:chemistry*
chemical_list: D005456:Fluorescent Dyes; D007473:Ion Channels; D008051:Lipid Bilayers; D008565:Membrane Proteins; D004247:DNA