Britton Chance Center for Biomedical Photonics, Wuhan National Laboratory for Optoelectronics-Huazhong University of Science and Technology, Wuhan, Hubei 430074, China.
MoE Key Laboratory for Biomedical Photonics, Collaborative Innovation Center for Biomedical Engineering, School of Engineering Sciences, Huazhong University of Science and Technology, Wuhan, Hubei 430074, China.
J Am Chem Soc. 2021 Feb 3;143(4):1917-1923. doi: 10.1021/jacs.0c10792. Epub 2021 Jan 20.
G-Quadruplex (G4) is a noncanonical nucleic acid secondary structure with multiple biofunctions. Identifying G4-related proteins (G4RPs) is important for understanding the roles of G4 in biology. Current methods to identify G4RPs include discovery from specific biological processes or pull-down assays with specific G4 sequences. Here, we report an strategy used to identify G4RPs with extensive sequence tolerance based on G4 ligand-mediated cross-linking. Applying this method, we identified 114 and 281 G4RPs in SV589 and MM231 cells, respectively. The results successfully overlapped with all the pull-down assay literature. Through the electrophoretic mobility shift assay (EMSA), we identified some new G4-binding proteins. Moreover, enhanced cross-linking and immunoprecipitation (eCLIP) confirmed that one newly identified G4-binding protein, SERBP1, interacts with G4 in the cellular environment. The method we developed provides a new strategy for identifying proteins that interact with nucleic secondary structures in cells and benefit the study of their biological roles.
四链体（G4）是一种具有多种生物功能的非经典核酸二级结构。鉴定与四链体相关的蛋白（G4RPs）对于理解 G4 在生物学中的作用很重要。目前鉴定 G4RPs 的方法包括从特定的生物学过程中发现或使用特定的 G4 序列进行下拉实验。在这里，我们报告了一种基于 G4 配体介导的交联来鉴定具有广泛序列容忍度的 G4RPs 的策略。应用该方法，我们分别在 SV589 和 MM231 细胞中鉴定到 114 个和 281 个 G4RPs。结果与所有下拉实验文献成功重叠。通过电泳迁移率变动分析（EMSA），我们鉴定了一些新的 G4 结合蛋白。此外，交联增强和免疫沉淀（eCLIP）证实，新鉴定的 G4 结合蛋白 SERBP1 在细胞环境中与 G4 相互作用。我们开发的方法为鉴定与细胞中核酸二级结构相互作用的蛋白质提供了一种新策略，有助于研究它们的生物学作用。
