title: Crystallization and preliminary X-ray analysis of a recombinant Fab fragment in complex with 17beta-oestradiol.
abstract: The recombinant Fab fragment of the anti-17beta-oestradiol antibody 57-2 has been a target for several protein-engineering experiments. A method for production, purification and crystallization of the Fab fragment alone (apo form) and in complex with the major female sex hormone 17beta-oestradiol is reported here. Diffracting apo-form crystals were only obtained with microseeding; crystals of the Fab-steroid complex were produced by co-crystallization in the presence of oestradiol and cross-seeding with the apo-form crystals. The crystals were grown using vapour-diffusion methods with reservoir solutions containing 10-14% PEG 4000 or 8-12% PEG 8000 and Tris-HCl buffer at high pH (9.0-9.5). Both the apo and complex crystals belong to space group P2(1)2(1)2(1) and diffract to 2.0 A resolution. High-resolution X-ray data sets suitable for structure determination were collected from flash-cooled crystals using 25% glycerol as the cryoprotectant.
authors.affiliation: Department of Biotechnology, University of Turku, Tykistönkatu 6, 6th floor, 20500 Turku, Finland. urpo.lamminmaki@utu.fi
authors.forename: U
authors.initials: U
authors.lastname: Lamminmäki
authors.forename: J
authors.initials: J
authors.lastname: Kankare
mesh_terms: D018360:Crystallography, X-Ray; D004926:Escherichia coli; D004958:Estradiol / Q000737:chemistry*; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification
chemical_list: D007140:Immunoglobulin Fab Fragments; D011994:Recombinant Proteins; D004958:Estradiol