NUCLEOSOME POSITIONING IS DETERMINED BY THE (H3-H4)2 TETRAMER
NUCLEOSOME POSITIONING IS DETERMINED BY THE (H3-H4)2 TETRAMER
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DOI：
10.1073/pnas.88.23.10596
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发表时间：
1991-12-01
影响因子：
11.1
通讯作者：
VANHOLDE, KE
中科院分区：
文献类型：
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作者：
DONG, F;VANHOLDE, KE
研究表明，即使在没有其他组蛋白的情况下，组蛋白(H3-H4)2四聚体也可以在DNA上找到特定的位置。用盐梯度透析法将纯化的组蛋白(H3-H4)2四聚体重组到含有核小体定位序列的208个碱基对的DNA分子上。组蛋白四聚体与DNA的化学计量比为1：1。用微球菌核酸酶消化后，可形成大小约为73bp的受保护DNA片段。用限制性内切酶切割73个碱基的DNA产生了一小组明确的条带，证明了(H3-H4)2四聚体在DNA上的定位。限制性内切酶分析表明，73bp的DNA主要对应于两个片段，一个位于该DNA上完整的组蛋白八聚体所采用的主要位置的伪二分体轴的两侧。这一结果意味着单个(H3-H4)2组蛋白四聚体可以折叠几乎等于-146个碱基的DNA，其定位与完整的八聚体相同，但在没有组蛋白H_2A和H_2B的情况下，伪二分体附近的区域仅能弱保护免受微球菌核酸酶的攻击。
It is demonstrated that the histone (H3-H4)2 tetramer can find specific positions on DNA, even in the absence of other histones. Purified histone (H3-H4)2 tetramers were reconstituted onto 208-base-pair (bp) DNA molecules containing a nucleosome-positioning sequence by using salt-gradient dialysis. The stoichiometry of histone tetramer to DNA was shown to be 1:1. Digestion with micrococcal nuclease led to formation of protected DNA fragments of almost-equal-to 73 bp. Cleavage of the 73-bp DNA with restriction enzymes produced a small set of defined bands, demonstrating positioning of the (H3-H4)2 tetramer on DNA. Analysis of the restriction digests shows that the 73-bp DNA corresponds mainly to two fragments, one, lying on either side of the pseudo-dyad axis of the major position adopted by complete histone octamers on this DNA. This result means that a single (H3-H4)2 histone tetramer can fold almost-equal-to 146 bp of DNA with the same positioning as the complete octamer but that a region near the pseudo-dyad is only weakly protected against micrococcal nuclease attack in the absence of histones H2A and H2B.