title: Combining dehydration, construct optimization and improved data collection to solve the crystal structure of a CRM1-RanGTP-SPN1-Nup214 quaternary nuclear export complex.
abstract: High conformational flexibility is an intrinsic and indispensable property of nuclear transport receptors, which makes crystallization and structure determination of macromolecular complexes containing exportins or importins particularly challenging. Here, the crystallization and structure determination of a quaternary nuclear export complex consisting of the exportin CRM1, the small GTPase Ran in its GTP-bound form, the export cargo SPN1 and an FG repeat-containing fragment of the nuclear pore complex component nucleoporin Nup214 fused to maltose-binding protein is reported. Optimization of constructs, seeding and the development of a sophisticated protocol including successive PEG-mediated crystal dehydration as well as additional post-mounting steps were essential to obtain well diffracting crystals.
authors.affiliation: Abteilung für Molekulare Strukturbiologie, Institut für Mikrobiologie und Genetik, Göttinger Zentrum für Molekulare Biowissenschaften (GZMB), Georg-August-Universität Göttingen, Justus-von-Liebig-Weg 11, 37077 Göttingen, Germany.
authors.forename: Thomas
authors.initials: T
authors.lastname: Monecke
authors.affiliation: Abteilung für Molekulare Strukturbiologie, Institut für Mikrobiologie und Genetik, Göttinger Zentrum für Molekulare Biowissenschaften (GZMB), Georg-August-Universität Göttingen, Justus-von-Liebig-Weg 11, 37077 Göttingen, Germany.
authors.forename: Achim
authors.initials: A
authors.lastname: Dickmanns
authors.affiliation: Macromolecular Crystallography (HZB-MX), Helmholtz-Zentrum Berlin für Materialien und Energie, Albert-Einstein-Strasse 15, 12489 Berlin, Germany.
authors.forename: Manfred S
authors.initials: MS
authors.lastname: Weiss
authors.affiliation: Institut für Molekularbiologie, Universitätsmedizin Göttingen, Georg-August-Universität Göttingen, Humboldtallee 23, 37073 Göttingen, Germany.
authors.forename: Sarah A
authors.initials: SA
authors.lastname: Port
authors.affiliation: Institut für Molekularbiologie, Universitätsmedizin Göttingen, Georg-August-Universität Göttingen, Humboldtallee 23, 37073 Göttingen, Germany.
authors.forename: Ralph H
authors.initials: RH
authors.lastname: Kehlenbach
authors.affiliation: Abteilung für Molekulare Strukturbiologie, Institut für Mikrobiologie und Genetik, Göttinger Zentrum für Molekulare Biowissenschaften (GZMB), Georg-August-Universität Göttingen, Justus-von-Liebig-Weg 11, 37077 Göttingen, Germany.
authors.forename: Ralf
authors.initials: R
authors.lastname: Ficner
mesh_terms: D021581:Active Transport, Cell Nucleus; D002467:Cell Nucleus / Q000378:metabolism*; D018360:Crystallography, X-Ray; D003890:Desiccation; D028884:Karyopherins / Q000737:chemistry*; D008958:Models, Molecular; D028861:Nuclear Pore Complex Proteins / Q000737:chemistry*; D018160:Receptors, Cytoplasmic and Nuclear / Q000737:chemistry*; D020931:ran GTP-Binding Protein / Q000737:chemistry*; D000097606:Exportin 1 Protein
chemical_list: D028884:Karyopherins; D028861:Nuclear Pore Complex Proteins; D018160:Receptors, Cytoplasmic and Nuclear; D020931:ran GTP-Binding Protein; D000097606:Exportin 1 Protein
keywords: CRM1; HC1c crystal humidifier; crystal dehydration; maltose-binding protein; nucleoporin