TY - JOUR
T1 - Protein expression, characterization, crystallization and preliminary X-ray crystallographic analysis of a Fic protein from Clostridium difficile
AU - Welner, Ditte
AU - Dedic, Emil
AU - van Leeuwen, Hans C.
AU - Kuijper, Ed
AU - Bjerrum, Morten Jannik
AU - Østergaard, Ole
AU - Jørgensen, Rene
PY - 2014/6
Y1 - 2014/6
N2 - 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%.
AB - 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%.
U2 - 10.1107/S2053230X1400987X
DO - 10.1107/S2053230X1400987X
M3 - Journal article
C2 - 24915103
SN - 1744-3091
VL - 70
SP - 827
EP - 831
JO - Acta Crystallographica. Section F: Structural Biology and Crystallization Communications Online
JF - Acta Crystallographica. Section F: Structural Biology and Crystallization Communications Online
ER -