Patterton H G, Hapgood J
Department of Biochemistry, University of Cape Town, South Africa.
Nucleic Acids Res. 1996 Nov 1;24(21):4349-55. doi: 10.1093/nar/24.21.4349.
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.
海胆G串珠结合因子（suGF1）是几种能与寡聚（dGxdC）基序序列特异性结合的蛋白质之一，该基序常出现在真核基因上游。在本研究中，我们研究了纯化至近乎均一的suGF1与其在体外重构核小体核心中的寡聚（dGxdC）结合位点之间的相互作用。我们发现，体外重构一个包含suGF1结合位点的214 bp片段会产生五种不同的核小体核心种类。这些种类的组蛋白八聚体具有相同的旋转取向，但平移框架不同。结果表明，suGF1结合位点在五种核心种类中的不同核小体定位会调节suGF1与核小体DNA结合的能力，尽管核小体核心中DNA的旋转取向最大程度地暴露了suGF1结合位点。我们提出，suGF1与组蛋白八聚体之间直接的蛋白质 - 蛋白质空间冲突最有可能是调节suGF1与其核小体包裹的结合位点结合的决定因素。这一结果表明，在体内，suGF1可能像TBP、NF1和热休克因子一样，需要一种互补的核小体破坏活性，或者suGF1在核小体沉积之前就与游离的新生复制DNA结合。
