DNA helicase activity in purified human RECQL4 protein.
Human RECQL4 protein was expressed in insect cells using a baculovirus protein expression system and it was purified to near homogeneity. The protein sedimented at a position between catalase (230 kDa) and ferritin (440 kDa) in glycerol gradient centrifugation, suggesting that it forms homo-multimers. Activity to displace annealed 17-mer oligonucleotide in the presence of ATP was co-sedimented with hRECQL4 protein. In ion-exchange chromatography, both DNA helicase activity and single-stranded DNA-dependent ATPase activity were co-eluted with hRECQL4 protein. The requirements of ATP and Mg for the helicase activity were different from those for the ATPase activity. The data suggest that the helicase migrates on single-stranded DNA in a 3'-5' direction. These results suggest that the hRECQL4 protein exhibits DNA helicase activity.
Suzuki
Takahiro
T
Ibaraki University, 2-1-1 Bunkyo, Mito, Ibaraki 310-8512, Japan.
Kohno
Toshiyuki
T
Ishimi
Yukio
Y
D000251:Adenosine Triphosphatases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000528:radiation effects; D000255:Adenosine Triphosphate / Q000378:metabolism / Q000528:radiation effects; D016367:Baculoviridae / Q000235:genetics; D002499:Centrifugation, Density Gradient; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D004265:DNA Helicases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism* / Q000528:radiation effects; D004256:DNA Polymerase I / Q000378:metabolism / Q000528:radiation effects; D018076:DNA, Complementary; D006367:HeLa Cells; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008274:Magnesium / Q000378:metabolism; D011089:Polydeoxyribonucleotides / Q000378:metabolism; D011485:Protein Binding; D053484:RecQ Helicases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism* / Q000528:radiation effects; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000528:radiation effects; D014466:Ultraviolet Rays
D018076:DNA, Complementary; D011089:Polydeoxyribonucleotides; D011994:Recombinant Proteins; D000255:Adenosine Triphosphate; D004256:DNA Polymerase I; D000251:Adenosine Triphosphatases; C506731:RECQL4 protein, human; C037693:single stranded DNA dependent ATPase; D004265:DNA Helicases; D053484:RecQ Helicases; D008274:Magnesium