Crystallization and preliminary X-ray diffraction studies of NusG, a protein shared by the transcription and translation machines.
N-utilization factor G (NusG) from Aquifex aeolicus (Aa) was overexpressed in Escherichia coli, purified and crystallized using the hanging-drop vapor-diffusion technique. The drops consisted of 2.5 microl protein solution (approximately 30 mg ml(-1) in 20 mM Tris-HCl pH 8.0, 200 mM NaCl, 2 mM EDTA and 10 mM DTT) and 2.5 microl reservoir solution (0.085 M Na HEPES pH 7.5, 15% glycerol, 11% 2-propanol and 20% PEG 4000) derived from condition number 41 of the Hampton Cryo Screen. The crystals grew at 291 +/- 1 K and reached dimensions of 0.2 x 0.1 x 0.05 mm in 5-7 d. The crystals, which diffracted to 2.45 A resolution, belonged to space group C222(1), with unit-cell parameters a = 65.95, b = 124.58, c = 83.60 A. One AaNusG molecule is present in the asymmetric unit, corresponding to a solvent content of 59.80% (Matthews coefficient = 3.06 A(3) Da(-1)). Crystal structure determination is in progress.
Andrykovitch
Michelle
M
Biomolecular Structure Section, Macromolecular Crystallography Laboratory, National Cancer Institute, National Institutes of Health, Frederick, MD 21702, USA.
Guo
Wei
W
Routzahn
Karen M
KM
Gu
Yijun
Y
Anderson
D Eric
DE
Reshetnikova
Ludmila S
LS
Knowlton
J Randolph
JR
Waugh
David S
DS
Ji
Xinhua
X
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D029968:Escherichia coli Proteins; D010445:Peptide Elongation Factors / Q000737:chemistry* / Q000235:genetics; D014176:Protein Biosynthesis; D011487:Protein Conformation; D014157:Transcription Factors / Q000737:chemistry* / Q000235:genetics; D014158:Transcription, Genetic
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; C070291:NusG protein, E coli; D010445:Peptide Elongation Factors; D014157:Transcription Factors