A ligand discovery toolbox for the WWE domain family of human E3 ligases

Journal article

The WWE domain is a relatively under-researched domain found in twelve human proteins and characterized by a conserved tryptophan-tryptophan-glutamate (WWE) sequence motif. Six of these WWE domain-containing proteins also contain domains with E3 ubiquitin ligase activity. The general recognition of poly-ADP-ribosylated substrates by WWE domains suggests a potential avenue for development of Proteolysis-Targeting Chimeras (PROTACs). Here, we present novel crystal structures of the HUWE1, TRIP12, and DTX1 WWE domains in complex with PAR building blocks and their analogs, thus enabling a comprehensive analysis of the PAR binding site structural diversity. Furthermore, we introduce a versatile toolbox of biophysical and biochemical assays for the discovery and characterization of novel WWE domain binders, including fluorescence polarization-based PAR binding and displacement assays, 15N-NMR-based binding affinity assays and 19F-NMR-based competition assays. Through these assays, we have characterized the binding of monomeric iso-ADP-ribose (iso-ADPr) and its nucleotide analogs with the aforementioned WWE proteins. Finally, we have utilized the assay toolbox to screen a small molecule fragment library leading to the successful discovery of novel ligands targeting the HUWE1 WWE domain.

Authors: Münzker, L; Kimani, SW; Fowkes, MM; Dong, A; Zheng, H

• Publication status: Published
• Peer review status: Peer reviewed
• Publisher: Nature Research
• Journal: Communications Biology More from this journal
• Volume: 7
• Issue: 1
• Article number: 901
• Publication date: 2024-07-24
• Acceptance date: 2024-07-14
• DOI: 10.1038/s42003-024-06584-w
• EISSN: 2399-3642
• ISSN: 2399-3642
• Language: English