SUBUNIT STRUCTURE OF CHROMATIN

这项研究的总体目标是了解 和相互作用，决定和稳定 核小体和染色质的高级结构。自.以来 这些结构中的可逆修改必须伴随着 染色质复制和转录等核过程，以及 了解染色质的稳定性对于一个完整的 理解这些过程是如何发生的以及如何在 活着。 这些问题将通过使用定义明确的 模型系统。核小体和寡核小体将是 组蛋白八聚体在克隆DNA上的重组构建 含有单个或多个定位序列的片段。 完整的鸡红细胞组蛋白或其来源的组蛋白 N末端的尾巴已经通过受控的蛋白水解会被移除 被雇佣。新方法允许在修改后的任何一种情况下进行重建 H_2A/H_2B或H_3/H_4。 利用这些系统，将调查以下问题： (1)哪些组蛋白或组蛋白的哪些部分识别 在DNA序列上定位信号？ (2)解离的热力学参数是什么？ 在中等盐浓度下有一个确定的核小体？什么是 特定组蛋白和部分组蛋白对 核小体稳定吗？ (3)扩展链的确切要求是什么 染色质中的螺线管转变？这些实验将利用 如上所述构建的阶段性均相寡核小体。 (4)不同类别的组蛋白尾巴在其中扮演什么角色 稳定螺线管结构？

The overall aim of this research is an understanding of the forces and interactions that determine and stabilize the structure of the nucleosome and the higher order structure of chromatin. Since reversible modification in these structures must accompany such nuclear processes as chromatin replication and transcription, an understanding of chromatin stability is essential to a complete comprehension of how these processes occur and are controlled in vivo. The problems will be approached through the use of well defined model systems. Nucleosomes and oligonucleosomes will be constructed by reconstitution with histone octamers on cloned DNA fragments containing single or multiple positioning sequences. Either intact chicken erythrocyte histones or histones from which N-terminal tails have been removed by controlled proteolysis will be employed. New methods allow reconstitution with either modified H2A/H2B or H3/H4. With such systems, the following questions will be investigated: (1) What histones, or portions of histones, recognize the positioning signals on DNA sequences? (2) What are the thermodynamic parameters for the dissociation of a defined nucleosome at moderate salt concentrations? What are the contributions of particular histones and portions of histones to nucleosome stability? (3) What are the precise requirements for the extended chain to solenoid transition in chromatin? These experiments will utilize phased, homogeneous oligonucleosomes constructed as above. (4) What roles do the different classes of histone tails play in stabilizing the solenoidal structure?