Protein-Protein and Protein-DNA Interfaces of the Nucleosome

本研究的目的是阐明控制核小体结构与细胞周期中发生的功能重塑（组装和拆卸之间的过渡）之间关系的特性。该方法是基于经典生物化学和生物物理化学与分子遗传学的综合应用。这种方法解决了核小体DNA和八聚体组蛋白核心在动态结构中的基本特性。来自PI实验室的新证据（基于分子动力学计算）表明核小体是非常动态的，在纳秒时间尺度上表现出巨大的“呼吸”运动，蛋白质亚基之间的接触界面显示出随时间的波动。蛋白质核心是一个“铰接的内骨骼”，DNA以“一般的，非碱基特异性的方式”包裹着它的大部分长度，相对较少的位点特异性接触。这种“脆弱”的安排很好地满足了监管的需要；它在很大程度上是由核心组蛋白八聚体内的亚基接触介导的。这些研究正在研究影响天然异源二聚体的内在（分子类型）和外在（微环境）参数及其组装潜力的一系列条件。这一点很重要，因为对核小体/染色质在染色质重塑、复制和转录过程中所涉及的功能转变的透彻理解取决于对这些界面接触的能量学和特异性的确切了解。将采用的技术有：分析超离心、高分辨率多角度光散射和微量热法以及当代分析化学方法。

Moudrianakis0091736The goal of the this research is to elucidate the properties that govern the relations between the structure of the nucleosome and the functional remodeling (transitions between assembly and disassembly) that occurs during the cell cycle. The approach is based on integrated applications of classical biochemistry and biophysical chemistry with those of molecular genetics. This approach addresses the basic properties of both nucleosomal DNA and the octameric histone core within a dynamic structure. New evidence from the PI's laboratory (based on molecular dynamics calculations) suggests that the nucleosome is very dynamic, exhibiting large "breathing" motions in the nanosecond time scale and that the contact interfaces between the protein subunits show time-dependent fluctuations. The protein core is an "articulated endoskeleton" around which the DNA wraps in a "generic, non base-specific way" for most of its length, with relatively few site-specific contacts. Such a "fragile" arrangement serves the needs of regulation well; it is mediated to a large extent by the subunit contacts within the core histone octamer. These studies are examining a range of conditions that affect the intrinsic (molecular types) and extrinsic (microenvironment) parameters of native heterodimers and their assembly potential. This is important because a thorough understanding of the functional transitions of the nucleosome/chromatin entailed during chromatin remodeling, replication and transcription depends on the exact knowledge of the energetics and specificities of these interfacial contacts. Techniques to be employed are: analytical ultracentrifugation, high-resolution multi-angle light scattering and microcalorimetry as well as contemporary methods of analytical chemistry.