Molecular Biophysics Unit, Indian Institute of Science, Bengaluru, India.
Protein Sci. 2022 May;31(5):e4294. doi: 10.1002/pro.4294.
BAF250b and its paralog BAF250a are the DNA-binding central hub proteins present in BAF-B and BAF-A classes of SWI/SNF chromatin-remodeling complexes. BAF250b contains an AT-rich interaction domain (ARID) and C-terminal BAF250_C domain, and it is found mutated in several cancers. ARID is a conserved helix-turn-helix motif-containing DNA-binding domain present in several eukaryotic proteins. The ARID of BAF250b has been proposed to play roles in recruiting SWI/SNF to the target gene promoters for their activation. BAF250b ARID structures had been deposited in the protein data bank by a structural genomics consortium. However, it is not well-studied for its DNA-binding and solution dynamic properties. Here, we report complete backbone NMR resonance assignments of human BAF250b ARID. NMR chemical shifts and the backbone dynamics showed that the solution structure of the protein matched the reported crystal structures. The structure and chemical shift indexing revealed the presence of a short β-sheet in the DNA-binding region of BAF250b ARID that was absent in the structure of its paralog BAF250a ARID. NMR chemical shift perturbations identified DNA-binding residues and revealed the DNA-binding interface on BAF250b ARID. NMR data-driven HADDOCK models of BAF250b ARID - DNA complexes revealed its plausible mode of DNA-binding. Isothermal titration calorimetry experiments showed that BAF250b ARID interacts with DNA sequences with moderate affinities like BAF250a ARID. However, distinct thermodynamic signatures were observed for binding of BAF250a ARID and BAF250b ARID to AT-rich DNA sequence, suggesting that subtle sequence and structural differences in these two proteins influence their DNA-binding.
BAF250b 及其同源物 BAF250a 是存在于 BAF-B 和 BAF-A 类 SWI/SNF 染色质重塑复合物中的 DNA 结合中心蛋白。BAF250b 包含富含 AT 的相互作用结构域（ARID）和 C 端 BAF250_C 结构域，并且在几种癌症中发现其发生突变。ARID 是一个保守的螺旋-转角-螺旋结构域，存在于几种真核蛋白中。BAF250b 的 ARID 被认为在招募 SWI/SNF 到靶基因启动子以激活它们的过程中发挥作用。BAF250b ARID 的结构已被结构基因组学联盟存入蛋白质数据库。然而，其 DNA 结合和溶液动态特性尚未得到很好的研究。在这里，我们报告了人 BAF250b ARID 的完整的骨架 NMR 共振归属。NMR 化学位移和骨架动力学表明，该蛋白质的溶液结构与报道的晶体结构相匹配。结构和化学位移索引揭示了 BAF250b ARID 的 DNA 结合区存在一个短的 β-折叠，而其同源物 BAF250a ARID 的结构中则不存在。NMR 化学位移扰动鉴定了 DNA 结合残基，并揭示了 BAF250b ARID 的 DNA 结合界面。基于 NMR 数据的 HADDOCK 模型显示了 BAF250b ARID 与 DNA 复合物的可能结合模式。等温滴定量热实验表明，BAF250b ARID 与 DNA 序列以中等亲和力相互作用，类似于 BAF250a ARID。然而，BAF250a ARID 和 BAF250b ARID 与富含 AT 的 DNA 序列的结合表现出不同的热力学特征，这表明这两种蛋白质在序列和结构上的细微差异影响它们的 DNA 结合。
