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Chemical profiling of DNA G-quadruplex-interacting proteins in live cells

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Balasubramanian, Shankar  ORCID logo
Adhikari, Santosh 
Martinez Cuesta, Sergio 


DNA-protein interactions regulate critical biological processes. Identifying proteins that bind to specific, functional genomic loci is essential for understanding the underlying regulatory mechanisms on a molecular level. Here, we describe a co-binding-mediated protein profiling (CMPP) strategy to investigate the interactome of DNA G-quadruplexes (G4s) in native chromatin. CMPP involves cell-permeable, functionalized G4-ligand probes that bind endogenous G4s and subsequently crosslink to co-binding G4-interacting proteins in situ. We first show the robustness of CMPP by proximity labelling of a G4 binding protein in vitro. Employing this approach in live cells, we then identify hundreds of putative G4-interacting proteins from various functional classes. Next, we confirmed high G4 binding affinity and selectivity for several newly discovered G4 interactors in vitro, and we validate direct G4 interactions for a functionally important candidate in cellular chromatin using an independent approach. Our studies provide a chemical strategy to map protein interactions of specific nucleic acid features in living cells.



Alkynes, Aminoquinolines, Cell Line, Tumor, Cross-Linking Reagents, DNA, DNA-Binding Proteins, Diazomethane, G-Quadruplexes, HEK293 Cells, Humans, Ligands, Proof of Concept Study, Protein Binding, Ultraviolet Rays

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Nature Chemistry

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Nature Research
Cancer Research UK (CB4330)
Cancer Research UK (C9681/A29214)
Wellcome Trust (209441/Z/17/Z)
Herchel Smith funds