ArticleChemical science2024
High-throughput assay exploiting disorder-to-order conformational switches: application to the proteasomal Rpn10:E6AP complex.
Article in Chemical science, 2024. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 2 papers.
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Who cites it
2 citing papers in PubMed, 2 citations in OpenAlex.
- NMR in the Age of Modern Biomedical Research and Drug Discovery.Journal of molecular biology · 2025Review
- Optimized isolation of enzymatically active ubiquitin E3 ligase E6AP/UBE3A from mammalian cells.Protein expression and purification · 2025Article
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
Abstract
Conformational switching is pervasively driven by protein interactions, particularly for intrinsically disordered binding partners. We developed a dually orthogonal fluorescence-based assay to monitor such events, exploiting environmentally sensitive fluorophores. This assay is applied to E3 ligase E6AP, as its AZUL domain induces a disorder-to-order switch in an intrinsically disordered region of the proteasome, the so-named Rpn10 AZUL-binding domain (RAZUL). By testing various fluorophores, we developed an assay appropriate for high-throughput screening of Rpn10:E6AP-disrupting ligands. We found distinct positions in RAZUL for fluorophore labeling with either acrylodan or Atto610, which had disparate spectral responses to E6AP binding. E6AP caused a hypsochromic shift with increased fluorescence of acrylodan-RAZUL while decreasing fluorescence intensity of Atto610-RAZUL. Combining RAZUL labeled with either acrylodan or Atto610 into a common sample achieved robust and orthogonal measurement of the E6AP-induced conformational switch. This approach is generally applicable to disorder-to-order (or
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Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.