Bionanoscience Department, Kavli Institute of Nanoscience, Delft University of Technology, Van der Maasweg 9, 2629 HZ Delft, The Netherlands.
Division of Clinical Biochemistry, Biocenter, Medical University of Innsbruck, Innrain 80, 6020 Innsbruck, Austria.
J Chem Phys. 2018 Mar 28;148(12):123323. doi: 10.1063/1.5009100.
Nucleosomes consisting of a short piece of deoxyribonucleic acid (DNA) wrapped around an octamer of histone proteins form the fundamental unit of chromatin in eukaryotes. Their role in DNA compaction comes with regulatory functions that impact essential genomic processes such as replication, transcription, and repair. The assembly of nucleosomes obeys a precise pathway in which tetramers of histones H3 and H4 bind to the DNA first to form tetrasomes, and two dimers of histones H2A and H2B are subsequently incorporated to complete the complex. As viable intermediates, we previously showed that tetrasomes can spontaneously flip between a left-handed and right-handed conformation of DNA-wrapping. To pinpoint the underlying mechanism, here we investigated the role of the H3-H3 interface for tetramer flexibility in the flipping process at the single-molecule level. Using freely orbiting magnetic tweezers, we studied the assembly and structural dynamics of individual tetrasomes modified at the cysteines close to this interaction interface by iodoacetamide (IA) in real time. While such modification did not affect the structural properties of the tetrasomes, it caused a 3-fold change in their flipping kinetics. The results indicate that the IA-modification enhances the conformational plasticity of tetrasomes. Our findings suggest that subnucleosomal dynamics may be employed by chromatin as an intrinsic and adjustable mechanism to regulate DNA supercoiling.
核小体由一段短的脱氧核糖核酸（DNA）缠绕在组蛋白八聚体周围组成，是真核生物染色质的基本单位。它们在 DNA 紧缩中的作用伴随着调节功能，影响着复制、转录和修复等基本基因组过程。核小体的组装遵循一个精确的途径，其中组蛋白 H3 和 H4 的四聚体首先与 DNA 结合形成四聚体，然后两个组蛋白 H2A 和 H2B 的二聚体被随后并入以完成复合物。作为可行的中间体，我们之前表明四聚体可以在 DNA 缠绕的左手和右手构象之间自发翻转。为了确定潜在的机制，我们在这里研究了 H3-H3 界面在四聚体灵活性翻转过程中的作用在单分子水平上。使用自由旋转的磁镊，我们实时研究了在靠近该相互作用界面的半胱氨酸处用碘乙酰胺（IA）修饰的单个四聚体的组装和结构动力学。虽然这种修饰不影响四聚体的结构特性，但会导致其翻转动力学发生 3 倍变化。结果表明，IA 修饰增强了四聚体的构象可塑性。我们的发现表明亚核小体动力学可能被染色质用作内在和可调机制来调节 DNA 超螺旋。
