title: Generation of recombinant antibody fragments for membrane protein crystallization.
abstract: Membrane proteins are challenging targets for crystallization and structure determination by X-ray crystallography. Hurdles can be overcome by antibody-mediated crystallization. More than 25 unique structures of membrane protein:antibody complexes have already been determined. In the majority of cases, hybridoma-derived antibody fragments either in Fab or Fv fragment format were employed for these complexes. We will briefly introduce the background and current status of the strategy and describe in detail the current protocols of well-established methods for the immunization, the selection, and the characterization of antibodies, as well as the cloning, the production, and the purification of recombinant antibodies useful for structural analysis of membrane proteins.
authors.affiliation: Institute for Biochemistry and Molecular Biology, ZBMZ, BIOSS Centre for Biological Signalling Studies, University of Freiburg, Freiburg, Germany; Department of Clinical Biochemistry, University of Kashmir, Srinagar, India.
authors.forename: Syed H
authors.initials: SH
authors.lastname: Mir
authors.affiliation: Institute for Biochemistry and Molecular Biology, ZBMZ, BIOSS Centre for Biological Signalling Studies, University of Freiburg, Freiburg, Germany.
authors.forename: Claudia
authors.initials: C
authors.lastname: Escher
authors.affiliation: Institute for Biochemistry and Molecular Biology, ZBMZ, BIOSS Centre for Biological Signalling Studies, University of Freiburg, Freiburg, Germany.
authors.forename: Wei-Chun
authors.initials: WC
authors.lastname: Kao
authors.affiliation: Institute for Biochemistry and Molecular Biology, ZBMZ, BIOSS Centre for Biological Signalling Studies, University of Freiburg, Freiburg, Germany; Faculty of Biology, University of Freiburg, Freiburg, Germany.
authors.forename: Dominic
authors.initials: D
authors.lastname: Birth
authors.affiliation: Institute for Biochemistry and Molecular Biology, ZBMZ, BIOSS Centre for Biological Signalling Studies, University of Freiburg, Freiburg, Germany.
authors.forename: Christophe
authors.initials: C
authors.lastname: Wirth
authors.affiliation: Institute for Biochemistry and Molecular Biology, ZBMZ, BIOSS Centre for Biological Signalling Studies, University of Freiburg, Freiburg, Germany. Electronic address: carola.hunte@biochemie.uni-freiburg.de.
authors.forename: Carola
authors.initials: C
authors.lastname: Hunte
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D003001:Cloning, Molecular / Q000379:methods*; D003460:Crystallization / Q000379:methods; D018360:Crystallography, X-Ray / Q000379:methods; D004926:Escherichia coli / Q000235:genetics; D007114:Immunization; D007128:Immunoglobulin Fragments / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D008807:Mice, Inbred BALB C; D010843:Pichia / Q000235:genetics; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification
chemical_list: D000911:Antibodies, Monoclonal; D007128:Immunoglobulin Fragments; D008565:Membrane Proteins; D011994:Recombinant Proteins
keywords: Antibodies; Crystallization; Fab fragment; Fv fragment; Membrane proteins; X-ray structures