Das Tulika, Mukhopadhyay Swagata, Das Amit K, Ganguly Agneyo
Department of Bioscience and Biotechnology, Indian Institute of Technology Kharagpur, Kharagpur 721302, West Bengal, India.
Nucleic Acids Res. 2025 Jun 6;53(11). doi: 10.1093/nar/gkaf489.
RecQ helicases can unwind a wide spectrum of DNA structures and thereby protect the cells from genome instability. Unwinding mechanisms have been extensively studied for bacterial and human RecQ helicases. DNA-induced winged helix (WH) domain repositioning and allosteric remodeling of the ATPase domain have been shown to be important for unwinding activity of bacterial RecQ helicases. In contrast, no such altered conformational state was observed for human RECQ1 upon DNA or nucleotide binding. In this study, we have crystallized and characterized an engineered RECQ1 containing a flexible glycine serine-rich linker inserted between the zinc binding and WH domains. The linker containing construct exhibits more efficient DNA binding and unwinding activity. A crystal structure of the engineered RECQ1 in complex with DNA exhibits conformational rearrangements of the helicase and WH domains, resulting in a more compact structure. Our structure, for the first time, demonstrates that alteration of the distance between the tip of the β-hairpin and the ARL favors DNA binding and remodels the ATPase domain, leading to alteration in substrate recognition and unwinding activity. These structural rearrangements are necessary for efficient coordination between the WH domain and the helicase domain, coupling DNA binding and ATP hydrolysis to strand separation.
RecQ解旋酶能够解开多种DNA结构，从而保护细胞免受基因组不稳定的影响。对于细菌和人类RecQ解旋酶的解旋机制已经进行了广泛研究。已证明DNA诱导的翼状螺旋（WH）结构域重新定位和ATP酶结构域的变构重塑对于细菌RecQ解旋酶的解旋活性很重要。相比之下，在结合DNA或核苷酸时，未观察到人类RECQ1有这种构象状态的改变。在本研究中，我们结晶并表征了一种经过工程改造的RECQ1，它在锌结合结构域和WH结构域之间插入了一个富含甘氨酸丝氨酸的柔性接头。含有接头的构建体表现出更有效的DNA结合和解旋活性。与DNA复合的工程化RECQ1的晶体结构显示了解旋酶和WH结构域的构象重排，形成了更紧凑的结构。我们的结构首次证明，β-发夹末端与ARL之间距离的改变有利于DNA结合并重塑ATP酶结构域，导致底物识别和解旋活性的改变。这些结构重排对于WH结构域和解旋酶结构域之间的有效协调是必要的，将DNA结合和ATP水解与链分离偶联起来。
