Folding mechanism of the (H3-H4)2 histone tetramer of the core nucleosome
Folding mechanism of the (H3-H4)2 histone tetramer of the core nucleosome
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DOI：
10.1110/ps.03535504
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发表时间：
2004-05-01
期刊：
影响因子：
8
通讯作者：
Gloss, LM
中科院分区：
文献类型：
--
作者：
Banks, DD;Gloss, LM
为了进一步理解寡聚蛋白质组装，已经检查了H3-H4历史四聚体的折叠和解折叠动力学。四聚体是核心核小体的核心蛋白组分，核心核小体是真核细胞核中DNA压缩成染色质的基本单位。这份报告提供了第一个动力学折叠研究的蛋白质含有历史性的折叠二聚化基序，基序中观察到的几种蛋白质-DNA复合物。先前的平衡解折叠研究已经证明，在生理条件下，在H3-H4二聚体和四聚体种类之间存在动态平衡。在有机渗透剂三甲胺-N-氧化物（TMAO）的存在下，这种平衡主要向四聚体移动。停流方法，监测固有的酪氨酸荧光和远紫外圆二色性，已被用来测量折叠和展开动力学作为盐酸胍（盐酸钆）和单体浓度的函数，在0和IM TMAO。动力学相的分配通过使用H3突变体C110 E对专性H3-H4二聚体的研究来辅助，所述突变体C110 E使H3-H3'疏水性四螺旋束四聚体界面不稳定。H3-H4体系的动力学折叠机制是一个顺序过程。未折叠的H3和H4单体在爆发相反应中缔合以形成二聚体中间体，其经历进一步的一级折叠过程以在折叠途径的限速步骤中形成天然二聚体。然后，H3-H4二聚体以107 M（-1）sec（-1）的速率常数快速缔合，以在完全组装的四聚体和折叠的H3-H4二聚体之间建立动态平衡。
To further understand oligomeric protein assembly, the folding and unfolding kinetics of the H3-H4 historic tetramer have been examined. The tetramer is the central protein component of the core nucleosome, which is the basic unit of DNA compaction into chromatin in the eukaryotic nucleus. This report provides the first kinetic folding studies of a protein containing the historic fold dimerization motif, a motif observed in Several protein-DNA complexes. Previous equilibrium unfolding studies have demonstrated that, under physiological conditions, there is a dynamic equilibrium between the H3-H4 dimer and tetramer species. This equilibrium is shifted predominantly toward the tetramer in the presence of the organic osmolyte trimethylamine-N-oxide (TMAO). Stopped-flow methods, monitoring intrinsic tyrosine fluorescence and far-UV circular dichroism, have been used to measure folding and unfolding kinetics as a function of guanidinium hydrochloride (GdnHCl) and monomer concentrations, in 0 and I M TMAO. The assignment of the kinetic phases was aided by the study of an obligate H3-H4 dimer, using the H3 mutant, C110E, which destabilizes the H3-H3' hydrophobic four-helix bundle tetramer interface. The proposed kinetic folding mechanism of the H3-H4 system is a sequential process. Unfolded H3 and H4 monomers associate in a burst phase reaction to form a dimeric intermediate that undergoes a further, first-order folding process to form the native dimer in the rate-limiting step of the folding pathway. H3-H4 dimers then rapidly associate with a rate constant of 107 M(-1)sec(-1) to establish a dynamic equilibrium between the fully assembled tetramer and folded H3-H4 dimers.