Chemoproteomic profiling to identify activity changes and functional inhibitors of DNA-binding proteins. Issue 11 (17th November 2022)

• Record Type: Journal Article
• Authors: Ruprecht, Benjamin; Wei, Lan; Zheng, Li; Bodea, Smaranda; Mo, Xuan; Maschberger, Melanie; Stoehr, Gabriele; Hahne, Hannes; Cornella-Taracido, Ivan; Chi, An
• Is Part Of: Cell chemical biology. Volume 29:Issue 11(2022)
• Page Start: 1639
• Page End: 1648.e4
• Publication Date: 2022-11-17
• Subjects: chemical proteomics -- mass spectrometry -- DNA binding -- drug screening -- reactive side chain profiling -- pull-down -- covalency; Biochemistry -- Periodicals; 572.05
• DOI: 10.1016/j.chembiol.2022.10.008
• Languages: English
• ISSNs: 2451-9456

Abstract

Summary: DNA-binding proteins are promising therapeutic targets but are notoriously difficult to drug. Here, we evaluate a chemoproteomic DNA interaction platform as a complementary strategy for parallelized compound profiling. To enable this approach, we determined the proteomic binding landscape of 92 immobilized DNA sequences. Perturbation-induced activity changes of captured transcription factors in disease-relevant settings demonstrated functional relevance of the enriched subproteome. Chemoproteomic profiling of >300 cysteine-directed compounds against a coverage optimized bead mixture, which specifically captures >150 DNA binders, revealed competition of several DNA-binding proteins, including the transcription factors ELF1 and ELF2. We also discovered the first compound that displaces the DNA-repair complex MSH2-MSH3 from DNA. Compound binding to cysteine 252 on MSH3 was confirmed using chemoproteomic reactive cysteine profiling. Overall, these results suggested that chemoproteomic DNA bead pull-downs enable the specific readout of transcription factor activity and can identify functional "hotspots" on DNA binders toward expanding the druggable proteome. Graphical abstract: Highlights: Proteomic characterization of native protein complexes bound to >90 DNA sequences Assembly of a bead mixture to specifically enrich >150 native DNA binders Quantification of compound-induced protein competition from DNA bead mixture Identification of the first compound that disrupts MSH2-MSH3 binding to DNA Abstract : Ruprecht et al. applied mass-spectrometry-based proteomics to characterize the native protein binding landscape of >90 immobilized DNA sequences. The multiplexed chemoproteomic enrichment of active protein complexes from disease-relevant lysate enabled the parallelized identification of functional chemical disruptors for DNA binders and transcription factors.