Overexpression, purification, crystallization and preliminary X-ray diffraction analysis of the pMV158-encoded plasmid transcriptional repressor protein CopG.
Plasmid pMV158 encodes a 45 amino acid transcriptional repressor, CopG, which is involved in copy number control. A new procedure for overproduction and purification of the protein has been developed. The CopG protein thus obtained retained its ability to specifically bind to DNA and to repress its own promoter. Purified CopG protein has been crystallized using the sitting-drop vapor diffusion method. The crystals, belonging to orthorhombic space group C222(1) (cell constants a = 67.2 A, b = 102.5 A, c = 40.2 A), were obtained from a solution containing methylpentanediol, benzamidine and sodium chloride, buffered to pH 6.7. Complete diffraction data up to 1.6 A resolution have been collected. Considerations about the Matthews parameter account for the most likely presence of three molecules in the asymmetric unit (2.27 A3/Da).
Gomis-Rüth
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FX
Centre d'Investigació i Desenvolupament, C.S.I.C., Barcelona, Spain. xgrcri@cid.csic.es
Solà
M
M
Pérez-Luque
R
R
Acebo
P
P
Alda
M T
MT
González
A
A
Espinosa
M
M
del Solar
G
G
Coll
M
M
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D018360:Crystallography, X-Ray; D004265:DNA Helicases; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D010957:Plasmids; D017433:Protein Structure, Secondary; D011506:Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification; D015534:Trans-Activators; D014961:X-Ray Diffraction
D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D011506:Proteins; D011994:Recombinant Proteins; D012097:Repressor Proteins; D015534:Trans-Activators; C045264:replication initiator protein; D004265:DNA Helicases