Cai Sheng, Zhu Lingyang, Zhang Ziming, Chen Yuan
Division of Immunology, Beckman Research Institute of the City of Hope, 1500 Duarte Road, Duarte, California 91006, USA.
Biochemistry. 2007 May 1;46(17):4943-50. doi: 10.1021/bi061738h. Epub 2007 Apr 4.
Understanding the mechanism of protein-DNA interactions at the molecular level is one of the main focuses in structural and molecular biological investigations. At present, NMR spectroscopy is the only approach that can provide atomic details of protein-DNA recognition in solution. However, determining the structures of protein-DNA complexes using NMR spectroscopy has been dependent on the observation of intermolecular nuclear Overhauser effects (NOE) and their assignments, which are difficult to obtain in many cases. In this study, we have shown that intermolecular distance constraints derived from a single spin-label in combination with docking calculations have defined many specific contacts of the complex between the AT-rich interaction domain (ARID) of Mrf2 and its target DNA. Mrf2 contacts DNA mainly using the two flexible loops, L1 and L2. While the L1 loop contacts the phosphate backbone, L2 and several residues in the adjacent helices interact with AT base pairs in the major groove of DNA. Despite the structural diversity in the ARID family of DNA-binding proteins, Mrf2 maintains contacts with DNA similar to those observed in the homologous Dri-DNA complex.
在分子水平上理解蛋白质与DNA相互作用的机制是结构生物学和分子生物学研究的主要重点之一。目前，核磁共振光谱法是唯一能够提供溶液中蛋白质与DNA识别原子细节的方法。然而，利用核磁共振光谱法确定蛋白质-DNA复合物的结构一直依赖于分子间核Overhauser效应（NOE）及其归属的观察，而在许多情况下这些很难获得。在本研究中，我们表明，源自单个自旋标记的分子间距离限制与对接计算相结合，确定了Mrf2富含AT的相互作用结构域（ARID）与其靶DNA之间复合物的许多特定接触。Mrf2主要利用两个柔性环L1和L2与DNA接触。L1环与磷酸骨架接触，而L2环和相邻螺旋中的几个残基与DNA大沟中的AT碱基对相互作用。尽管DNA结合蛋白的ARID家族存在结构多样性，但Mrf2与DNA的接触方式与同源Dri-DNA复合物中观察到的相似。
