EWS是RNA/DNA结合蛋白家族FET的成员，调节核酸代谢的重要阶段。EWS包含一个N端低复杂度区域(LCD)和一个C端RNA结合域(RBD)。RBD进一步分为三个RG丰富区域，环绕一个RNA识别蛋白(RRM)和一个锌指(ZnF)域。最近，在Ewing肉瘤中，EWS被证明可以调节R-环，这是一种儿童骨骼和软组织癌症，在这种肿瘤中，染色体易位将EWS的N端LCD与转录因子FLI1的C端DNA结合域融合在一起。虽然已经证明EWS直接结合了R-环，但结合机制尚未阐明。在本研究中，EWS的RBD被分成几个构建块，随后对预计在R-环中形成的各种核酸结构进行了结合测试，包括RNA茎环、DNA G四联体和RNA:DNA杂合物。EWS与这三种不同的核酸结构相互作用，亲和力不同，多个结构域对于结合每种底物都有贡献。RRM和RG2区域似乎会混杂地结合核酸，而ZnF对单链结构显示出更强的选择性。通过这些结果，更好地理解了EWS对R-环和其他核酸结构的识别和结合的结构基础。 © 2023 Wiley Periodicals LLC.

EWS is a member of the FET family of RNA/DNA binding proteins that regulate crucial phases of nucleic acid metabolism. EWS comprises an N-terminal low-complexity domain (LCD) and a C-terminal RNA-binding domain (RBD). The RBD is further divided into three RG-rich regions, which flank an RNA-recognition motif (RRM) and a zinc finger (ZnF) domain. Recently, EWS was shown to regulate R-loops in Ewing sarcoma, a pediatric bone and soft-tissue cancer in which a chromosomal translocation fuses the N-terminal LCD of EWS to the C-terminal DNA binding domain of the transcription factor FLI1. Though EWS was shown to directly bind R-loops, the binding mechanism was not elucidated. In the current study, the RBD of EWS was divided into several constructs, which were subsequently assayed for binding to various nucleic acid structures expected to form at R-loops, including RNA stem-loops, DNA G-quadruplexes, and RNA:DNA hybrids. EWS interacted with all three nucleic acid structures with varying affinities and multiple domains contributed to binding each substrate. The RRM and RG2 region appear to bind nucleic acids promiscuously while the ZnF displayed more selectivity for single-stranded structures. With these results, the structural underpinnings of EWS recognition and binding of R-loops and other nucleic acid structures is better understood.© 2023 Wiley Periodicals LLC.