Unwinding of nucleosomal DNA by a DNA helicase.
We have asked whether a DNA helicase can unwind DNA contained within both isolated native chromatin and reconstituted chromatin containing regularly spaced arrays of nucleosome cores on a linear tandem repeat sequence. We find that Escherichia coli recBCD enzyme is capable of unwinding these DNA substrates and displacing the nucleosomes, although both the rate and the processivity of enzymatic unwinding are inhibited (a maximum of 3- and > 25-fold, respectively) as the nucleosome density on the template is increased. The observed rate of unwinding is not affected if the histone octamer is chemically cross-linked; thus, dissociation, or splitting, of the histone octamer is not required for unwinding to occur. The unwinding of native chromatin isolated from HeLa cell nuclei occurs both in the absence and in the presence of linker histone H1. These results suggest that as helicases unwind DNA, they facilitate nuclear processes by acting to clear DNA of histones or DNA-binding proteins in general.
Eggleston
A K
AK
Section of Microbiology, University of California, Davis 95616.
O'Neill
T E
TE
Bradbury
E M
EM
Kowalczykowski
S C
SC
D000818:Animals; D002843:Chromatin / Q000378:metabolism*; D004265:DNA Helicases / Q000378:metabolism*; D004926:Escherichia coli / Q000201:enzymology; D043211:Exodeoxyribonuclease V; D005090:Exodeoxyribonucleases / Q000378:metabolism*; D006367:HeLa Cells; D006657:Histones / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D009707:Nucleosomes / Q000378:metabolism*; D012617:Sea Urchins
D002843:Chromatin; D006657:Histones; D009707:Nucleosomes; D005090:Exodeoxyribonucleases; D043211:Exodeoxyribonuclease V; D004265:DNA Helicases