The translational placement of nucleosome cores in vitro determines the access of the transacting factor suGF1 to DNA.
The sea urchin G-string binding factor (suGF1) is one of several proteins that bind sequence-specifically to oligo(dGxdC) motifs, frequently present upstream of eukaryotic genes. In this study we investigate the interaction of suGF1, purified to near homogeneity, with its oligo(dGxdC) binding site in a reconstituted nucleosome core in vitro. We show that the in vitro reconstitution of a 214 bp fragment containing a suGF1 binding site results in the appearance of five distinct nucleosome core species. These species contain the histone octamer in an identical rotational setting but in different translational frames. The resulting different nucleosomal locations of the suGF1 binding site in the five core species are shown to modulate the ability of suGF1 to bind to nucleosomal DNA, even though the rotational setting of the DNA in the nucleosome cores maximally exposes the suGF1 binding site. We propose that a direct protein-protein steric clash between suGF1 and the histone octamer is the most likely determinant in modulating the binding of suGF1 to its nucleosomally wrapped binding site. This result suggests that in vivo suGF1, like TBP, NF1 and heat shock factor, may require a complementary nucleosome disrupting activity or that suGF1 binds to free nascent replicated DNA prior to nucleosome deposition.
Patterton
H G
HG
Department of Biochemistry, University of Cape Town, South Africa.
Hapgood
J
J
D000818:Animals; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D006657:Histones / Q000235:genetics; D009687:Nuclear Proteins / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D009707:Nucleosomes / Q000378:metabolism*; D009838:Oligodeoxyribonucleotides / Q000378:metabolism; D011485:Protein Binding; D014176:Protein Biosynthesis; D011487:Protein Conformation; D012617:Sea Urchins; D015534:Trans-Activators / Q000378:metabolism*
D004268:DNA-Binding Proteins; D006657:Histones; D009687:Nuclear Proteins; D009707:Nucleosomes; D009838:Oligodeoxyribonucleotides; D015534:Trans-Activators; C085454:suGF1 protein, Parechinus angulosus; C052201:oligo(dG-dC); D004247:DNA