高迁移率组 (HMG) 蛋白是固有无序的核非组蛋白染色体蛋白,通过调节真核细胞中众多基因的表达,在许多生物过程中发挥重要作用。HMGA 蛋白包含三个 DNA 结合基序,即“AT 钩”,它们优先与 B 型 DNA 小沟中富含 AT 的序列结合。从医学的角度来看,了解 AT 钩结构域与 DNA 的相互作用非常重要,因为 HMGA 蛋白参与不同的疾病,包括癌症和寄生虫病。我们在这里介绍了 HMGA AT 钩 1 结构域的第一个晶体结构(1.40 Å 分辨率),它与富含 AT 的 DNA 的小沟结合。与使 DNA 弯曲并显示更大的小沟变宽的 AT 钩 3 相反,AT-钩 1 结合相邻的 DNA 分子并在结合时显示 DNA 适度变宽。用表面等离子体共振 (SPR) 生物传感器和等温滴定量热法 (ITC) 实验在溶液中研究结合亲和力和结合的热力学。AT 钩 1 与含有 (TTAA)2 的 DNA 形成熵驱动的 2:1 复合物,具有相对较慢的缔合/解离动力学。我们表明 N-苯基苯甲酰胺衍生的抗动质体化合物 (1-3) 强烈且特异性地与 AT-DNA 的小沟结合,并与 AT 钩 1 竞争结合。分子的中心核心是观察到这些化合物序列选择性的基础。这些发现提供了关于与主要被忽视的热带病(如利什曼病和锥虫病)相关的 DNA 小沟结合化合物可能的作用方式的线索。
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Crystal structure of the HMGA AT-hook 1 domain bound to the minor groove of AT-rich DNA and inhibition by antikinetoplastid drugs
High mobility group (HMG) proteins are intrinsically disordered nuclear non-histone chromosomal proteins that play an essential role in many biological processes by regulating the expression of numerous genes in eukaryote cells. HMGA proteins contain three DNA binding motifs, the “AT-hooks”, that bind preferentially to AT-rich sequences in the minor groove of B-form DNA. Understanding the interactions of AT-hook domains with DNA is very relevant from a medical point of view because HMGA proteins are involved in different conditions including cancer and parasitic diseases. We present here the first crystal structure (1.40 Å resolution) of the HMGA AT-hook 1 domain, bound to the minor groove of AT-rich DNA. In contrast to AT-hook 3 which bends DNA and shows a larger minor groove widening, AT-hook 1 binds neighbouring DNA molecules and displays moderate widening of DNA upon binding. The binding affinity and thermodynamics of binding were studied in solution with surface plasmon resonance (SPR)-biosensor and isothermal titration calorimetry (ITC) experiments. AT-hook 1 forms an entropy-driven 2:1 complex with (TTAA)2-containing DNA with relatively slow kinetics of association/dissociation. We show that N-phenylbenzamide-derived antikinetoplastid compounds (1–3) bind strongly and specifically to the minor groove of AT-DNA and compete with AT-hook 1 for binding. The central core of the molecule is the basis for the observed sequence selectivity of these compounds. These findings provide clues regarding a possible mode of action of DNA minor groove binding compounds that are relevant to major neglected tropical diseases such as leishmaniasis and trypanosomiasis.