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Structural biology of transcriptional regulation by SATB1 that binds to matrix attachment region of DNA

SATB1 与 DNA 基质附着区结合的转录调控的结构生物学

基本信息

批准号：
17570103
负责人：
YAMASAKI Kazuhiko
金额：
$ 1.73万
依托单位：
National Institute of Advanced Industrial Science and Technology (AIST)
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

项目摘要

SATB1 (special AT-rich sequence binding protein 1) is a transcription factor that regulates maturation of immune T-cells in thymus. It represses transcription of several genes including interleukin 2 receptor by binding to the matrix attachment region (MAR) of DNA of the target gene promoters and by recruiting histone deacetylase to the binding site. In this study we revealed the mechanism of recognition of MAR-DNA by SATB1 by means of structural biology.We first determined the structure of MAR binding domain (MBD) of SATB1 by NMR spectroscopy. The structure possesses five a-helices arranged in a novel topology. By NMR titration experiments and surface plasmon resonance analyses of DNA binding of mutant proteins, the region of the protein relevant to DNA binding was identified. Also, by surface plasmon resonance experiments with DNA including modified bases and groove-specific binding drugs, it was revealed that SATB1-MBD binds to DNA from the major groove side, which is different from what was expected previously.Next, we crystallized the complex of SATB1-MBD and MAR-DNA reflecting up to 2.0 Å in the laboratory diffractometer or 1.75 Å at synchrotron. Data revealed a structure of the complex in which SATB1-MBD binds to the major groove of DNA. Between the molecules direct hydrogen bonds are observed between a single amino acid and a single base, and water-mediated hydrogen bonds and hydrophobic interactions play a major role in the sequence-specific recognition. SATB1-MBD belongs to CUT domain and this is the first report on the mechanism of DNA recognition by CUT domains.

SATB1是一种转录因子，调节胸腺免疫T细胞的成熟。它通过与靶基因启动子DNA的基质附着区(MAR)结合并在结合部位募集组蛋白脱乙酰酶来抑制包括白介素2受体在内的几个基因的转录。在本研究中，我们从结构生物学的角度揭示了SATB1识别MAR-DNA的机制。首先用核磁共振波谱确定了SATB1的MAR结合域(MBD)结构。该结构具有5个以新颖的拓扑结构排列的α-螺旋。通过核磁共振滴定实验和突变蛋白与DNA结合的表面等离子体共振分析，确定了与DNA结合相关的蛋白质区域。此外，通过对修饰碱基和沟槽特异性结合药物的表面等离子体共振实验，发现SATB_1-MBD从主槽一侧与DNA结合，这与之前的预期不同。其次，我们在实验室衍射仪上结晶了SATB_1-MBD与MAR-DNA的络合物，在实验室衍射仪上结晶了反射高达2.0Å的MAR-DNA，在同步加速器下结晶了反射高达1.75Å的SATB_1-MBD与MAR-DNA的复合物。数据揭示了SATB1-MBD与DNA主沟结合的复合体的结构。在分子之间，观察到单个氨基酸和单个碱基之间的直接氢键，水介导的氢键和疏水相互作用在序列特异性识别中起主要作用。SATB1-MBD属于CUT结构域，这是关于CUT结构域识别DNA机制的首次报道。