Luchnik A N, Bakayev V V, Zbarsky I B, Georgiev G P
EMBO J. 1982;1(11):1353-8. doi: 10.1002/j.1460-2075.1982.tb01322.x.
After treatment of SV40 minichromosomes with DNA topoisomerase I, the superhelicity in the bulk of the DNA extracted from minichromosomes is known to remain unchanged. However, we found that the DNA extracted from a small fraction of SV40 minichromosomes (2-5%), was almost completely relaxed, and covalently closed as shown by agarose gel electrophoresis. Thus, the DNA in these 2-5% of SV40 minichromosomes was probably torsionally strained (TS). The proportion of such TS minichromosomes is close to the estimated proportion of transcriptionally active minichromosomes. The distribution of the TS minichromosomes in sucrose gradient coincided with the distribution of transcriptionally active complexes. Both sedimented faster than the majority of minichromosomes. Furthermore, after treatment with topoisomerase I the relaxed minichromosomes could be quantitatively separated from the bulk of material by recentrifugation in a sucrose gradient. A major part of the endogenous RNA polymerase activity was recovered in the relaxed fraction. These data suggest that TS-minichromosomes correspond to transcriptionally active chromatin. After relaxation with topoisomerase I the TS minichromosomes lacked histones.
用DNA拓扑异构酶I处理SV40微型染色体后，已知从微型染色体中提取的大部分DNA的超螺旋度保持不变。然而，我们发现从一小部分SV40微型染色体（2 - 5%）中提取的DNA几乎完全松弛，并如琼脂糖凝胶电泳所示为共价闭合。因此，这2 - 5%的SV40微型染色体中的DNA可能存在扭转应变（TS）。这种TS微型染色体的比例接近于转录活性微型染色体的估计比例。TS微型染色体在蔗糖梯度中的分布与转录活性复合物的分布一致。两者沉降速度均快于大多数微型染色体。此外，用拓扑异构酶I处理后，通过在蔗糖梯度中再次离心，可将松弛的微型染色体与大部分物质定量分离。大部分内源性RNA聚合酶活性在松弛部分中得以恢复。这些数据表明TS - 微型染色体对应于转录活性染色质。用拓扑异构酶I松弛后，TS微型染色体缺乏组蛋白。
