From the National Laboratory of Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China and.
J Biol Chem. 2014 Feb 21;289(8):4882-95. doi: 10.1074/jbc.M113.501940. Epub 2013 Dec 30.
Retinoblastoma-binding protein 1 (RBBP1) is a tumor and leukemia suppressor that binds both methylated histone tails and DNA. Our previous studies indicated that RBBP1 possesses a Tudor domain, which cannot bind histone marks. In order to clarify the function of the Tudor domain, the solution structure of the RBBP1 Tudor domain was determined by NMR and is presented here. Although the proteins are unrelated, the RBBP1 Tudor domain forms an interdigitated double Tudor structure similar to the Tudor domain of JMJD2A, which is an epigenetic mark reader. This indicates the functional diversity of Tudor domains. The RBBP1 Tudor domain structure has a significant area of positively charged surface, which reveals a capability of the RBBP1 Tudor domain to bind nucleic acids. NMR titration and isothermal titration calorimetry experiments indicate that the RBBP1 Tudor domain binds both double- and single-stranded DNA with an affinity of 10-100 μM; no apparent DNA sequence specificity was detected. The DNA binding mode and key interaction residues were analyzed in detail based on a model structure of the Tudor domain-dsDNA complex, built by HADDOCK docking using the NMR data. Electrostatic interactions mediate the binding of the Tudor domain with DNA, which is consistent with NMR experiments performed at high salt concentration. The DNA-binding residues are conserved in Tudor domains of the RBBP1 protein family, resulting in conservation of the DNA-binding function in the RBBP1 Tudor domains. Our results provide further insights into the structure and function of RBBP1.
视网膜母细胞瘤结合蛋白 1（RBBP1）是一种肿瘤和白血病抑制因子，可与甲基化组蛋白尾巴和 DNA 结合。我们之前的研究表明，RBBP1 具有一个不能结合组蛋白标记的结构域。为了阐明 Tudor 结构域的功能，我们通过 NMR 确定了 RBBP1 Tudor 结构域的溶液结构，并在此呈现。尽管这些蛋白质之间没有关系，但 RBBP1 Tudor 结构域形成类似于组蛋白去甲基化酶 JMJD2A 的双 Tudor 结构域的交错双 Tudor 结构，后者是一种表观遗传标记读取器。这表明 Tudor 结构域具有功能多样性。RBBP1 Tudor 结构域具有显著的正电荷表面区域，这揭示了 RBBP1 Tudor 结构域结合核酸的能力。NMR 滴定和等温滴定量热法实验表明，RBBP1 Tudor 结构域以 10-100 μM 的亲和力结合双链和单链 DNA；没有检测到明显的 DNA 序列特异性。根据通过使用 NMR 数据的 HADDOCK 对接构建的 Tudor 结构域-dsDNA 复合物模型结构，详细分析了 DNA 结合模式和关键相互作用残基。静电相互作用介导了 Tudor 结构域与 DNA 的结合，这与在高盐浓度下进行的 NMR 实验一致。DNA 结合残基在 RBBP1 蛋白家族的 Tudor 结构域中保守，导致 RBBP1 Tudor 结构域的 DNA 结合功能保守。我们的结果为 RBBP1 的结构和功能提供了进一步的见解。
