title: Genetic code expansion for multiprotein complex engineering.
abstract: We present a baculovirus-based protein engineering method that enables site-specific introduction of unique functionalities in a eukaryotic protein complex recombinantly produced in insect cells. We demonstrate the versatility of this efficient and robust protein production platform, 'MultiBacTAG', (i) for the fluorescent labeling of target proteins and biologics using click chemistries, (ii) for glycoengineering of antibodies, and (iii) for structure-function studies of novel eukaryotic complexes using single-molecule Förster resonance energy transfer as well as site-specific crosslinking strategies.
authors.affiliation: European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
authors.forename: Christine
authors.initials: C
authors.lastname: Koehler
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Organic Chemistry, Karlsruhe, Germany.
authors.forename: Paul F
authors.initials: PF
authors.lastname: Sauter
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Organic Chemistry, Karlsruhe, Germany.
authors.forename: Mirella
authors.initials: M
authors.lastname: Wawryszyn
authors.affiliation: European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
authors.forename: Gemma Estrada
authors.initials: GE
authors.lastname: Girona
authors.affiliation: European Molecular Biology Laboratory (EMBL), Grenoble, Grenoble, France.
authors.forename: Kapil
authors.initials: K
authors.lastname: Gupta
authors.affiliation: European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
authors.forename: Jonathan J M
authors.initials: JJ
authors.lastname: Landry
authors.affiliation: European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
authors.forename: Markus Hsi-Yang
authors.initials: MH
authors.lastname: Fritz
authors.affiliation: European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
authors.forename: Ksenija
authors.initials: K
authors.lastname: Radic
authors.affiliation: European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
authors.forename: Jan-Erik
authors.identifier: http://orcid.org/0000-0003-1444-2136
authors.initials: JE
authors.lastname: Hoffmann
authors.affiliation: Wellcome Trust Centre for Cell Biology, Institute of Cell Biology, School of Biological Sciences, University of Edinburgh, Edinburgh, United Kingdom.
authors.forename: Zhuo A
authors.initials: ZA
authors.lastname: Chen
authors.affiliation: Wellcome Trust Centre for Cell Biology, Institute of Cell Biology, School of Biological Sciences, University of Edinburgh, Edinburgh, United Kingdom.
authors.forename: Juan
authors.initials: J
authors.lastname: Zou
authors.affiliation: European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
authors.forename: Piau Siong
authors.initials: PS
authors.lastname: Tan
authors.affiliation: Vienna Biocenter Core Facilities (VBCF GmbH), Vienna, Austria.
authors.forename: Bence
authors.identifier: http://orcid.org/0000-0003-1464-3034
authors.initials: B
authors.lastname: Galik
authors.affiliation: Vienna Biocenter Core Facilities (VBCF GmbH), Vienna, Austria.
authors.forename: Sini
authors.identifier: http://orcid.org/0000-0003-3754-5584
authors.initials: S
authors.lastname: Junttila
authors.affiliation: Vienna Biocenter Core Facilities (VBCF GmbH), Vienna, Austria.
authors.forename: Peggy
authors.initials: P
authors.lastname: Stolt-Bergner
authors.affiliation: Division of Pathology and Laboratory Medicine, European Institute of Oncology, Milan, Italy.
authors.forename: Giancarlo
authors.initials: G
authors.lastname: Pruneri
authors.affiliation: Vienna Biocenter Core Facilities (VBCF GmbH), Vienna, Austria.
authors.forename: Attila
authors.initials: A
authors.lastname: Gyenesei
authors.affiliation: European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
authors.forename: Carsten
authors.initials: C
authors.lastname: Schultz
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Organic Chemistry, Karlsruhe, Germany.
authors.forename: Moritz Bosse
authors.initials: MB
authors.lastname: Biskup
authors.affiliation: European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
authors.forename: Hueseyin
authors.initials: H
authors.lastname: Besir
authors.affiliation: European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
authors.forename: Vladimir
authors.identifier: http://orcid.org/0000-0002-0352-2547
authors.initials: V
authors.lastname: Benes
authors.affiliation: Wellcome Trust Centre for Cell Biology, Institute of Cell Biology, School of Biological Sciences, University of Edinburgh, Edinburgh, United Kingdom.|Chair of Bioanalytics, Institute of Biotechnology, Technische Universität Berlin, Berlin, Germany.
authors.forename: Juri
authors.identifier: http://orcid.org/0000-0001-5999-1310
authors.initials: J
authors.lastname: Rappsilber
authors.affiliation: European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Jechlinger
authors.affiliation: European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
authors.forename: Jan O
authors.identifier: http://orcid.org/0000-0002-2798-3794
authors.initials: JO
authors.lastname: Korbel
authors.affiliation: European Molecular Biology Laboratory (EMBL), Grenoble, Grenoble, France.|The School of Biochemistry, University of Bristol, Bristol, United Kingdom.
authors.forename: Imre
authors.initials: I
authors.lastname: Berger
authors.affiliation: Karlsruhe Institute of Technology (KIT), Institute of Organic Chemistry, Karlsruhe, Germany.|Karlsruhe Institute of Technology (KIT), Institut für Toxikologie und Genetik, Eggenstein-Leopoldshafen, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Braese
authors.affiliation: European Molecular Biology Laboratory (EMBL), Heidelberg, Germany.
authors.forename: Edward A
authors.initials: EA
authors.lastname: Lemke
mesh_terms: D000818:Animals; D016367:Baculoviridae / Q000235:genetics / Q000378:metabolism; D018929:Cell Culture Techniques; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods; D005815:Genetic Code; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D006801:Humans; D046912:Multiprotein Complexes / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D010957:Plasmids; D015202:Protein Engineering / Q000379:methods*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D061987:Sf9 Cells; D018411:Spodoptera; D014764:Viral Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics
chemical_list: D046912:Multiprotein Complexes; D011994:Recombinant Proteins; D014764:Viral Proteins; D049452:Green Fluorescent Proteins