Crystallization and preliminary X-ray analyses of catabolite control protein A, free and in complex with its DNA-binding site.
The catabolite control protein (CcpA) from Bacillus megaterium is a member of the bacterial repressor protein family GalR/LacI. CcpA with an N-terminal His-tag was used for crystallization. Crystals of free CcpA and of CcpA in complex with the putative operator sequence (catabolite responsive elements, CRE) were obtained by vapour-diffusion techniques at 291 K using the hanging-drop method. CcpA crystals grown in the presence of polyethylene glycol 8000 belong to the hexagonal space group P6(1)22 or P6(5)22, with unit-cell parameters a = 74.4, c = 238.8 A. These crystals diffract X-rays to 2.55 A resolution and contain one monomer of the homodimeric protein per asymmetric unit. Crystals of the CcpA-CRE complex were obtained with ammonium sulfate as precipitant and belong to the tetragonal space group I4(1)22, with unit-cell parameters a = 125, c = 400 A and one complex per asymmetric unit. Although these co-crystals grew to a sufficient size, X-ray diffraction was limited to 8 A resolution.
Tebbe
J
J
Institut für Kristallographie, Freie Universität Berlin, Takustrasse 6, D-14195 Berlin, Germany.
Orth
P
P
Küster-Schöck
E K
EK
Hillen
W
W
Saenger
W
W
Hinrichs
W
W
D000595:Amino Acid Sequence; D001410:Bacillus megaterium / Q000737:chemistry / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D001483:Base Sequence; D001665:Binding Sites; D003460:Crystallization; D018360:Crystallography, X-Ray; D004269:DNA, Bacterial / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D004268:DNA-Binding Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*; D004926:Escherichia coli / Q000235:genetics; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification; D012097:Repressor Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D012097:Repressor Proteins; C069286:catabolite control proteins, bacteria