Retinoblastoma-binding protein 1 (RBBP1) is involved in gene regulation, epigenetic regulation, and disease processes. RBBP1 contains five domains with DNA-binding or histone-binding activities, but how RBBP1 specifically recognizes chromatin is still unknown. An AT-rich interaction domain (ARID) in RBBP1 was proposed to be the key region for DNA-binding and gene suppression. Here, we first determined the solution structure of a tandem PWWP-ARID domain mutant of RBBP1 after deletion of a long flexible acidic loop L12 in the ARID domain. NMR titration results indicated that the ARID domain interacts with DNA with no GC- or AT-rich preference. Surprisingly, we found that the loop L12 binds to the DNA-binding region of the ARID domain as a DNA mimic and inhibits DNA binding. The loop L12 can also bind weakly to the Tudor and chromobarrel domains of RBBP1, but binds more strongly to the DNA-binding region of the histone H2A-H2B heterodimer. Furthermore, both the loop L12 and DNA can enhance the binding of the chromobarrel domain to H3K4me3 and H4K20me3. Based on these results, we propose a model of chromatin recognition by RBBP1, which highlights the unexpected multiple key roles of the disordered acidic loop L12 in the specific binding of RBBP1 to chromatin.

视网膜母细胞瘤结合蛋白 1 (RBBP1) 参与基因调控、表观遗传调控和疾病过程。RBBP1 包含五个具有 DNA 结合或组蛋白结合活性的结构域，但 RBBP1 如何特异性识别染色质尚不清楚。RBBP1 中富含 AT 的相互作用域 (ARID) 被认为是 DNA 结合和基因抑制的关键区域。在这里，我们首先确定了 RBBP1 串联 PWWP-ARID 域突变体在 ARID 域中的长柔性酸性环 L12 缺失后的溶液结构。NMR 滴定结果表明 ARID 结构域与 DNA 相互作用，没有富含 GC 或 AT 的偏好。令人惊讶的是，我们发现环 L12 作为 DNA 模拟物与 ARID 域的 DNA 结合区结合并抑制 DNA 结合。L12环也可以与RBBP1的Tudor和chromobarrel结构域弱结合，但与组蛋白H2A-H2B异源二聚体的DNA结合区结合更强。此外，环 L12 和 DNA 都可以增强色桶结构域与 H3K4me3 和 H4K20me3 的结合。基于这些结果，我们提出了 RBBP1 识别染色质的模型，该模型突出了无序酸性环 L12 在 RBBP1 与染色质的特异性结合中出人意料的多重关键作用。