A protein binds to a satellite DNA repeat at three specific sites that would be brought into mutual proximity by DNA folding in the nucleosome

Using a generally applicable assay for specific DNA-binding proteins in crude extracts, we have detected and purified an HMG-like nuclear protein from African green monkey cells that preferentially binds to the 172 bp repeat of alpha-satellite DNA (alpha-DNA). DNAase I footprinting with the purified protein detects three specific binding sites (I-III) per alpha-DNA repeat. Site II is 145 bp (one core nucleosome length) from site III on the adjacent alpha-DNA repeat, while site I lies midway between sites II and III. In the alpha-nucleosome phasing frame corresponding with this arrangement, sites I-III would be brought into mutual proximity by DNA folding in the nucleosome. This phasing frame is identical with the preferred frame detected previously in isolated chromatin. Our results suggest that this new and abundant protein recognizes a family of short, related nucleotide sequences found not only in alpha-DNA but also throughout the genome, and that functions of this protein are mediated through its nucleosome-positioning activity. Such nucleosome-positioning proteins may underlie the sequence specificity of both nucleosome arrangements and higher order chromatin structures.