Institute of Structural Biology and Drug Design, Virginia Commonwealth University, Richmond, VA 23298-0035, USA.
Nucleic Acids Res. 2011 Aug;39(15):6741-52. doi: 10.1093/nar/gkr262. Epub 2011 Apr 29.
The epigenetic code of DNA methylation is interpreted chiefly by methyl cytosine binding domain (MBD) proteins which in turn recruit multiprotein co-repressor complexes. We previously isolated one such complex, MBD2-NuRD, from primary erythroid cells and have shown it contributes to embryonic/fetal β-type globin gene silencing during development. This complex has been implicated in silencing tumor suppressor genes in a variety of human tumor cell types. Here we present structural details of chicken MBD2 bound to a methylated DNA sequence from the ρ-globin promoter to which it binds in vivo and mediates developmental transcriptional silencing in normal erythroid cells. While previous studies have failed to show sequence specificity for MBD2 outside of the symmetric mCpG, we find that this domain binds in a single orientation on the ρ-globin target DNA sequence. Further, we show that the orientation and affinity depends on guanine immediately following the mCpG dinucleotide. Dynamic analyses show that DNA binding stabilizes the central β-sheet, while the N- and C-terminal regions of the protein maintain mobility. Taken together, these data lead to a model in which DNA binding stabilizes the MBD2 structure and that binding orientation and affinity is influenced by the DNA sequence surrounding the central mCpG.
DNA 甲基化的表观遗传密码主要由甲基胞嘧啶结合域（MBD）蛋白来解读，这些蛋白反过来又招募多蛋白共抑制复合物。我们之前从原代红细胞中分离出这样的一个复合物 MBD2-NuRD，并表明它有助于胚胎/胎儿β型珠蛋白基因在发育过程中的沉默。该复合物已被认为与多种人类肿瘤细胞类型中的肿瘤抑制基因沉默有关。在这里，我们展示了鸡 MBD2 与 ρ-珠蛋白启动子上的一段甲基化 DNA 序列结合的结构细节，该序列在体内与之结合，并在正常红细胞中介导发育转录沉默。虽然之前的研究未能在非对称 mCpG 之外显示 MBD2 的序列特异性，但我们发现该结构域以单一取向结合在 ρ-珠蛋白靶 DNA 序列上。此外，我们还表明，这种取向和亲和力取决于 mCpG 二核苷酸之后的鸟嘌呤。动态分析表明，DNA 结合稳定了中央β-折叠，而蛋白质的 N 端和 C 端区域保持了流动性。总之，这些数据表明 DNA 结合稳定了 MBD2 结构，而结合取向和亲和力受中央 mCpG 周围 DNA 序列的影响。
