Expression, purification and crystallization of the cell-division protein YgfE from Escherichia coli.
An open reading frame designated b2910 (ygfE) in the Escherichia coli K12-MG1655 genome sequence, identified as a possible homologue to the cell-division protein ZapA, was cloned into the high-expression plasmid pETDuet-1 and overexpressed in E. coli BL21 (DE3)-AI. The protein was purified in three steps to 99% purity. Crystals were obtained by the hanging-drop vapour-diffusion method at 291 K from a wide range of screened conditions, but principally from solutions containing 0.1 M HEPES pH 7.0, 18% PEG 6000, 5 mM CaCl2. Diffraction data to 1.8 A were collected at the European Synchrotron Radiation Facility (ESRF). The crystals belong to space group P6(1)22 or P6(5)22, with unit-cell parameters a = 53.8, b = 53.8, c = 329.7 A, alpha = beta = 90, gamma = 120 degrees.
Addinall
Stephen G
SG
Faculty of Life Sciences, University of Manchester, Oxford Road, Manchester M13 9PT, England.
Johnson
Kenneth A
KA
Dafforn
Timothy
T
Smith
Corinne
C
Rodger
Alison
A
Gomez
Raul Paco
RP
Sloan
Katherine
K
Blewett
Anne
A
Scott
David J
DJ
Roper
David I
DI
D002352:Carrier Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D002455:Cell Division; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000166:cytology / Q000201:enzymology; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification; D005784:Gene Amplification; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification
D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D011994:Recombinant Proteins; C520744:ZapA protein, E coli; C118060:ssnA protein, E coli