USE OF SELECTIVELY TRYPSINIZED NUCLEOSOME CORE PARTICLES TO ANALYZE THE ROLE OF THE HISTONE TAILS IN THE STABILIZATION OF THE NUCLEOSOME
USE OF SELECTIVELY TRYPSINIZED NUCLEOSOME CORE PARTICLES TO ANALYZE THE ROLE OF THE HISTONE TAILS IN THE STABILIZATION OF THE NUCLEOSOME
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DOI：
10.1016/0022-2836(89)90493-2
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发表时间：
1989-04-05
影响因子：
5.6
通讯作者：
VANHOLDE, KE
中科院分区：
文献类型：
--
作者：
AUSIO, J;DONG, F;VANHOLDE, KE
利用固定化胰蛋白酶和适当的分离程序，我们首次能够制备含有选择性胰蛋白酶化组蛋白结构域的核小体核心颗粒。由此得到的颗粒：[(H3T-H4T)2-2(H2At-H2BT)] .cntdot。DNA;[(H3-H4) 2 - 2 (H2AT-H2BT)] .cntdot。DNA;[H3T-H4T) 2 - 2 (H2A-H2B)] .cntdot。DNA（其中T表示胰蛋白酶化）以及非胰蛋白酶化的对照物已使用以下技术进行表征：分析性超离心、圆二色性、热变性和DNA酶I消化。本研究的主要目的是分析氨基末端区域（组蛋白“尾巴”）对溶液中核小体稳定性的作用。从该分析中获得的数据清楚地表明，核小体核心颗粒对解离的稳定性（低于0.7 M-NaCl的盐浓度）不受组蛋白n端区域存在或不存在的影响。此外，这些组蛋白区域对核小体在这种盐浓度范围内的构象转变的贡献很小，如果有的话。然而，它们在决定颗粒的热稳定性方面起着非常重要的作用，这反映在通过胰蛋白酶化选择性去除这些尾部时熔化轮廓所表现出的巨大变化中。熔化数据可以用一个简单的假设来解释，即将H2A/H2B和H3/H4尾部的相互作用归因于核小体DNA的特定区域。
Using immobilized trypsin and an appropriate fractionation procedure, we have been able to prepare, for the first time, nucleosome core particles containing selectively trypsinized histone domains. The particles thus obtained: [(H3T-H4T)2-2(H2At-H2BT)] .cntdot. DNA; [(H3-H4)2-2(H2AT-H2BT)] .cntdot. DNA; [H3T-H4T)2-2(H2A-H2B)] .cntdot. DNA (where T means trypsinized), together with the non-trypsinized controls have been characterized using the following techniques: analytical ultracentrifugation, circular dichroism, thermal denaturation and DNase I digestion. The major aim of this study was to analyze the role of the amino-terminal regions (the histone "tails") on the stability of the nucleosome in solution. The data obtained from this analysis clearly show that stability of the nucleosome core particle to dissociation (below a salt concentration of 0.7 M-NaCl) is not affected by the presence or the absence of any of the N-terminal regions of the histones. Furthermore, these histone regions make very little contribution, if any, to the conformational transition that nucleosomes undergo in this range of salt concentrations. They play, however, a very important role in determining the thermal stability of the particle, as reflected in the dramatic alterations exhibited by the melting profiles upon selective removal of these tails by trypsinization. The melting data can be explained by a simple hypothesis that ascribes interaction of H2A/H2B and H3/H4 tails to particular regions of the nucleosomal DNA.