Crystallization and preliminary X-ray crystallographic studies of the Z-DNA-binding domain of a PKR-like kinase (PKZ) in complex with Z-DNA.
PKZ, a PKR-like eIF2alpha kinase, consists of a Z-DNA-specific binding domain (Zalpha) and an eIF2alpha kinase domain. The kinase activity of PKZ is modulated by the binding of Zalpha to Z-DNA. The mechanisms underlying Z-DNA binding and the subsequent stimulation of PKZ raise intriguing questions. Interestingly, the Z-DNA-binding domain of PKZ from goldfish (Carassius auratus; caZalpha(PKZ)) shows limited sequence homology to other canonical Zalpha domains, suggesting that it may have a distinct Z-DNA-recognition mode. In this study, the Z-DNA-binding activity and stoichiometry of caZalpha(PKZ) were confirmed using circular dichroism (CD). In addition, preliminary X-ray studies of the caZalpha(PKZ)-Z-DNA complex are reported as the first step in the determination of its three-dimensional structure. Bacterially expressed recombinant caZalpha(PKZ) was purified and crystallized with Z-DNA at 295 K using the microbatch method. X-ray diffraction data were collected to 1.7 A resolution with an R(merge) of 7.4%. The crystals belonged to the monoclinic space group C2, with unit-cell parameters a = 55.54, b = 49.93, c = 29.44 A, beta = 96.5 degrees . Structural analysis of caZalpha(PKZ)-Z-DNA will reveal the binding mode of caZalpha(PKZ) to Z-DNA and its relevance to other Z-DNA-binding proteins.
Kim
Doyoun
D
Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon, Republic of Korea.
Hwang
Hye Yeon
HY
Kim
Yang Gyun
YG
Kim
Kyeong Kyu
KK
D000818:Animals; D002942:Circular Dichroism; D003460:Crystallization; D018360:Crystallography, X-Ray; D043542:DNA, Z-Form / Q000737:chemistry* / Q000378:metabolism*; D006054:Goldfish / Q000378:metabolism*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D019892:eIF-2 Kinase / Q000737:chemistry* / Q000378:metabolism*
D043542:DNA, Z-Form; D019892:eIF-2 Kinase