1. Expression, Biochemistry, and Stabilization with Camel Antibodies of Membrane Proteins: Case Study of the Mouse 5-HT3 Receptor
- Author
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Aline Desmyter, Horst Vogel, Lamia Mebarki, Takashi Tomizaki, Ruud Hovius, Romain Wyss, Sonja Minniberger, Xiao-Dan Li, Christophe Moreau, Lucie Peclinovska, Ghérici Hassaine, Cédric Deluz, Hugues Nury, Luigino Grasso, Henning Stahlberg, Laboratory of Physical Chemistry of Polymers and Membranes, Ecole Polytechnique Fédérale de Lausanne (EPFL), Biozentrum, University of Basel (Unibas), Architecture et fonction des macromolécules biologiques (AFMB), Institut National de la Recherche Agronomique (INRA)-Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), Institut de biologie structurale (IBS - UMR 5075), Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Grenoble Alpes (UGA)-Centre National de la Recherche Scientifique (CNRS), Laboratory of Biomolecular Research, Paul Scherrer Institute (PSI), Biozentrum [Basel, Suisse], Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)-Institut National de la Recherche Agronomique (INRA), Institut de biologie structurale (IBS - UMR 5075 ), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), and Lacapere, Jean-Jacques
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0301 basic medicine ,[SDV.BIO]Life Sciences [q-bio]/Biotechnology ,[SDV.NEU.NB]Life Sciences [q-bio]/Neurons and Cognition [q-bio.NC]/Neurobiology ,Gene Expression ,Crystallography, X-Ray ,[SDV.IMM.II]Life Sciences [q-bio]/Immunology/Innate immunity ,law.invention ,MESH: Recombinant Proteins ,Mice ,law ,[SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases ,MESH: Animals ,Integral membrane protein ,[SDV.MHEP.ME]Life Sciences [q-bio]/Human health and pathology/Emerging diseases ,biology ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM] ,Protein Stability ,Peripheral membrane protein ,MESH: Camelus ,[SDV.BIBS]Life Sciences [q-bio]/Quantitative Methods [q-bio.QM] ,Recombinant Proteins ,[SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM] ,[SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM] ,Biochemistry ,[SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology ,Recombinant DNA ,MESH: Membrane Proteins ,MESH: Cryoelectron Microscopy ,Antibody ,Camelus ,MESH: Gene Expression ,MESH: Receptors, Serotonin, 5-HT3 ,Cys-loop receptor ,VHH ,5-HT3 receptor ,Antibodies ,Cell Line ,03 medical and health sciences ,MESH: Protein Stability ,Animals ,Humans ,[SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM] ,MESH: Mice ,5-HT receptor ,MESH: Humans ,MESH: Antibodies ,Cryoelectron Microscopy ,Membrane Proteins ,MESH: Crystallography, X-Ray ,[SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology ,Lama antibody ,MESH: Cell Line ,030104 developmental biology ,Membrane protein ,Cell culture ,biology.protein ,[SDV.SP.PHARMA]Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology ,Receptors, Serotonin, 5-HT3 ,Stable cell line - Abstract
International audience; There is growing interest in the use of mammalian protein expression systems, and in the use of antibody-derived chaperones, for structural studies. Here, we describe protocols ranging from the production of recombinant membrane proteins in stable inducible cell lines to biophysical characterization of purified membrane proteins in complex with llama antibody domains. These protocols were used to solve the structure of the mouse 5-HT3 serotonin receptor but are of broad applicability for crystallization or cryo-electron microscopy projects.
- Published
- 2018
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