Crystallization and X-ray analysis of the N-terminal core domain of a tumour-associated human DEAD-box RNA helicase, rck/p54.
The RCK gene was cloned on the basis of the t(11;14)(q23;q32) chromosome translocation observed in human B-cell lymphoma cell line RC-K8. This gene was found to be overexpressed in various kinds of tumours. The gene product, rck/p54, consisting of 472 amino-acid residues with molecular weight 53.2 kDa, belongs to the family of DEAD-box RNA helicases. Its ATP-dependent RNA-unwinding activity toward c-myc RNA molecules in vitro has recently been demonstrated. In the present study, limited proteolysis experiments of rck/p54 were used to truncate the N-terminal domain (residues 1-288; 31.8 kDa) of rck/p54, leading to successful crystallization of Nc-rck/p54, i.e. the N-terminal core domain (residues 70-288; 24.5 kDa) of rck/p54. Crystals of Nc-rck/p54 were grown to a size suitable for X-ray structure analysis using polyethylene glycol 3350 as the precipitant. The crystal belongs to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 65.5, b = 73.1, c = 84.8 A, and diffracts X-rays to beyond 2.0 A resolution.
Matsui
Tsutomu
T
Department of Life Science, Tokyo Institute of Technology, 4259 Nagatsuta-cho, Midori-ku, Yokohama 226-8501, Japan.
Hogetsu
Keita
K
Akao
Yukihiro
Y
Tanaka
Masashi
M
Sato
Takao
T
Kumasaka
Takashi
T
Tanaka
Nobuo
N
D000595:Amino Acid Sequence; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D053487:DEAD-box RNA Helicases; D004247:DNA / Q000737:chemistry / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011518:Proto-Oncogene Proteins / Q000737:chemistry* / Q000235:genetics; D012316:RNA Nucleotidyltransferases / Q000737:chemistry* / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014357:Trypsin / Q000378:metabolism
D011518:Proto-Oncogene Proteins; D011994:Recombinant Proteins; D004247:DNA; D012316:RNA Nucleotidyltransferases; D014357:Trypsin; C094683:DDX6 protein, human; D053487:DEAD-box RNA Helicases