title: Protein expression, characterization, crystallization and preliminary X-ray crystallographic analysis of a Fic protein from Clostridium difficile.
abstract: Fic domains in proteins are found in abundance in nature from the simplest prokaryotes to animals. Interestingly, Fic domains found in two virulence factors of Gram-negative bacteria have recently been demonstrated to catalyse the transfer of the AMP moiety from ATP to small host GTPases. This post-translational modification has attracted considerable interest and a role for adenylylation in pathology and physiology is emerging. This work was aimed at the structural characterization of a newly identified Fic protein of the Gram-positive bacterium Clostridium difficile. A constitutively active inhibitory helix mutant of C. difficile Fic was overexpressed in Escherichia coli, purified and crystallized by the vapour-diffusion technique. Preliminary X-ray crystallographic analysis shows that the crystals diffract to at least 1.68 Å resolution at a synchrotron X-ray source. The crystals belonged to the orthorhombic space group P2₁2₁2₁, with unit-cell parameters a=45.6, b=80.8, c=144.7 Å, α=β=γ=90°. Two molecules per asymmetric unit corresponds to a Matthews coefficient of 2.37 Å3 Da(-1) and a solvent content of 48%.
authors.affiliation: Microbiology and Infection Control, Statens Serum Institut, Artillerivej 5, 2300 Copenhagen, Denmark.
authors.forename: Ditte
authors.initials: D
authors.lastname: Welner
authors.affiliation: Microbiology and Infection Control, Statens Serum Institut, Artillerivej 5, 2300 Copenhagen, Denmark.
authors.forename: Emil
authors.initials: E
authors.lastname: Dedic
authors.affiliation: Department of Medical Microbiology, Center of Infectious Diseases, Leiden University Medical Center, Albinusdreef 2, 2333 ZA Leiden, The Netherlands.
authors.forename: Hans C
authors.initials: HC
authors.lastname: van Leeuwen
authors.affiliation: Department of Medical Microbiology, Center of Infectious Diseases, Leiden University Medical Center, Albinusdreef 2, 2333 ZA Leiden, The Netherlands.
authors.forename: Ed
authors.initials: E
authors.lastname: Kuijper
authors.affiliation: Department of Chemistry, University of Copenhagen, Universitetsparken 5, 2100 Copenhagen, Denmark.
authors.forename: Morten Jannik
authors.initials: MJ
authors.lastname: Bjerrum
authors.affiliation: Department of Clinical Biochemistry and Immunology, Statens Serum Institut, Artillerivej 5, 2300 Copenhagen, Denmark.
authors.forename: Ole
authors.initials: O
authors.lastname: Østergaard
authors.affiliation: Microbiology and Infection Control, Statens Serum Institut, Artillerivej 5, 2300 Copenhagen, Denmark.
authors.forename: René
authors.initials: R
authors.lastname: Jørgensen
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D003001:Cloning, Molecular; D016360:Clostridioides difficile / Q000737:chemistry*; D003460:Crystallization; D018360:Crystallography, X-Ray / Q000379:methods*
chemical_list: D001426:Bacterial Proteins
keywords: Clostridium difficile; Fic protein; adenylylation