ARTIFICIAL NUCLEOSOME POSITIONING SEQUENCES
ARTIFICIAL NUCLEOSOME POSITIONING SEQUENCES
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DOI：
10.1073/pnas.86.19.7418
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发表时间：
1989-10-01
影响因子：
11.1
通讯作者：
CROTHERS, DM
中科院分区：
文献类型：
--
作者：
SHRADER, TE;CROTHERS, DM
我们已经使用了DNA弯曲性的序列依赖性的新兴规则来设计和测试一系列DNA分子，这些DNA分子强烈地并入核小体。竞争性重建实验表明，在组蛋白八聚体结合的DNA分子，其中部分完全由A和T或G和C，分开2个碱基对（bp），重复10 bp的周期的优势。这些重复的（A/T）3 NN（G/C）3 NN基序在核小体形成中比天然定位序列和其他重复基序如AANNNTTTNNN和GGNNNCNNN优越上级。不同长度的重复各向异性柔性DNA的研究表明，一段。嵌入160-bp片段中的40 bp足以产生与来自5S RNA基因的天然核小体定位序列相当的核小体结合。弯曲要求沿着表面的核小体似乎是相当恒定的，没有大的跳跃到结合自由能归因于蛋白质诱导的扭结。最有利的序列掺入到核小体中的强度比本体核小体DNA强100倍，但是当归一化为弯曲数时，微分弯曲自由能很小：每弯曲仅约100 cal/mol的自由能差（1 cal = 4.184 J）区分最佳弯曲序列和本体DNA。我们推断核小体中DNA弯曲的畸变能仅弱依赖于DNA序列。
We have used the emerging rules for the sequence dependence of DNA bendability to design and test a series of DNA molecules that incorporate strongly into nucleosomes. Competitive reconstitution experiments showed the superiority in histone octamer binding of DNA molecules in which segments consisting exclusively of A and T or G and c, separated by 2 base pairs (bp), are repeated with a 10-bp period. These repeated (A/T)3NN(G/C)3NN motifs are superior in nucleosome formation to natural positioning sequences and to other repeated motifs such as AANNNTTNNN and GGNNNCCNNN. Studies of different lengths of repetitive anisotropically flexible DNA showed that a segment of .apprxeq. 40 bp embedded in a 160-bp fragment is sufficient to generate nucleosome binding equivalent to that of natural nucleosome positioning sequences from 5S RNA genes. Bending requirements along the surface of the nucleosome seem to be quite constant, with no large jumps to binding free energy attributable to protein-induced kinks. The most favorable sequences incorporate into nucleosomes more strongly by 100-fold than bulk nucleosomal DNA, but differential bending free energies are small when normalized to the number of bends: a free energy difference of only about 100 cal/mol per bend (1 cal = 4.184 J) distinguishes the best bending sequences and bulk DNA. We infer that the distortion energy of DNA bending in the nucleosome is only weakly dependent on DNA sequence.