Use of a fusion protein to obtain crystals suitable for X-ray analysis: crystallization of a GST-fused protein containing the DNA-binding domain of DNA replication-related element-binding factor, DREF.
Crystals of glutathione-S-transferase (GST)-fused protein containing the DNA-binding domain of DNA replication-related element-binding factor, DREF, were obtained under crystallization conditions similar to those for GST. Preliminary X-ray crystallographic analysis revealed that crystals of the GST-fused protein belong to space group P6(1)22 or P6(5)22 with unit cell dimensions a = b = 140.4 A, c = 93.5 A and gamma = 120 degrees, having one molecule in the crystallographic asymmetric unit. The crystals diffract to 2.5 A resolution. The cell dimensions are related to those of GST crystals thus far reported. Crystallization of the DNA-binding domain that was cleaved from the fused protein by thrombin was also carried out using several methods under numerous conditions, but efforts to produce well-ordered large crystals were unsuccessful. A possible application of GST-fusion proteins for small target proteins or domains to obtain crystals suitable for X-ray structure determination is proposed.
Kuge
M
M
Department of Molecular Biology, Nara Institute of Science and Technology (NAIST), Japan.
Fujii
Y
Y
Shimizu
T
T
Hirose
F
F
Matsukage
A
A
Hakoshima
T
T
D018360:Crystallography, X-Ray; D004268:DNA-Binding Proteins / Q000737:chemistry*; D005982:Glutathione Transferase / Q000737:chemistry*; D011993:Recombinant Fusion Proteins / Q000737:chemistry*
D004268:DNA-Binding Proteins; D011993:Recombinant Fusion Proteins; D005982:Glutathione Transferase