title: Purification, crystallization and preliminary crystallographic analysis of DR0248, an MNT-HEPN fused protein from Deinococcus radiodurans.
abstract: DR0248 is a protein identified in the Deinococcus radiodurans (DR) genome that is predicted to encompass two domains: an N-terminal minimal nucleotidyl transferase domain (MNT) and a C-terminal higher eukaryotes and prokaryotes nucleotide-binding domain (HEPN). These two domains, usually encoded in two ORFs, have been suggested to play the role of a toxin-antitoxin (TA) system in prokaryotes. Recombinant DR0248 was overexpressed and purified from Escherichia coli and diffraction-quality crystals were obtained in the presence of the detergent molecules dodecyldimethylamine oxide (DDAO) and octaethylene glycol monododecyl ether (C12E8), which were used as crystallization additives. Crystals grown with DDAO diffracted to a resolution of 2.24 Å and belonged to space group C222(1), with unit-cell parameters a=98.4, b=129.9, c=59.2 Å. Crystals grown with C12E8 diffracted to a resolution of 1.83 Å and belonged to space group P2(1)2(1)2(1), with unit-cell parameters a=51.6, b=87.2, c=108.2 Å. The structure was solved by multiwavelength anomalous dispersion from zinc bound to the protein using a single crystal obtained in the presence of DDAO.
authors.affiliation: ESRF - The European Synchrotron, 71 Rue des Martyrs, Grenoble, France.
authors.forename: Gaelle
authors.initials: G
authors.lastname: Pesce
authors.affiliation: Departement de Biologie et de Genomique Structurales, Institute de Genetique et de Biologie Moleculaire et Cellulaire, 1 Rue Laurent Fries BP 10142, F-67400 Illkirch, France.
authors.forename: Simone
authors.initials: S
authors.lastname: Pellegrino
authors.affiliation: Department of Photon Science, Brookhaven National Laboratory, Upton, NY 11973-5000, USA.
authors.forename: Sean
authors.initials: S
authors.lastname: McSweeney
authors.affiliation: Macromolecular Crystallography Unit, Instituto de Tecnologia Quimica e Biologica, Universidade Nova de Lisboa, Oeiras, Portugal.
authors.forename: AnaMaria
authors.initials: A
authors.lastname: Goncalves
authors.affiliation: ESRF - The European Synchrotron, 71 Rue des Martyrs, Grenoble, France.
authors.forename: Daniele
authors.initials: D
authors.lastname: de Sanctis
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000302:isolation & purification; D020134:Catalytic Domain; D002850:Chromatography, Gel; D003460:Crystallization; D018360:Crystallography, X-Ray; D034301:Deinococcus; D009713:Nucleotidyltransferases / Q000737:chemistry* / Q000302:isolation & purification
chemical_list: D001426:Bacterial Proteins; D009713:Nucleotidyltransferases
keywords: DR0248; Deinococcus radiodurans; higher eukaryotes and prokaryotes nucleotide-binding domain; minimal nucleotidyl transferase domain