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Discovery of E6AP AZUL binding to UBQLN1/2 in cells, phase-separated droplets, and an AlphaFold-NMR integrated structure
- Publication Year :
- 2022
- Publisher :
- Cold Spring Harbor Laboratory, 2022.
-
Abstract
- The E3 ligase E6AP/UBE3A has a dedicated binding site in the 26S proteasome provided by the RAZUL domain of substrate receptor hRpn10/S5a/PSMD4. Guided by RAZUL sequence similarity, we test and demonstrate here that the E6AP AZUL binds transiently to the UBA of proteasomal shuttle factor UBQLN1/2. Despite a weak binding affinity, E6AP AZUL is recruited to UBQLN2 phase-separated droplets and E6AP interacts with UBQLN1/2 in cells. Steady-state and transfer NOE experiments indicate direct interaction of AZUL with the UBQLN1 UBA domain. Intermolecular contacts identified by NOESY data were combined with AlphaFold2-Multimer predictions to yield an AZUL:UBA model structure. We also identify a concentration-dependent oligomerization domain directly adjacent to UBQLN1/2 UBA (UBA-adjacent, UBAA) that is α-helical and allosterically reconfigured by AZUL binding to UBA. These data lead to a model of E6AP recruitment to UBQLN1/2 by AZUL:UBA interaction and provide fundamental information on binding requirements for interactions in droplets and cells.
Details
- Database :
- OpenAIRE
- Accession number :
- edsair.doi...........6161ef0fddcd0272d0357d2becd7b915
- Full Text :
- https://doi.org/10.1101/2022.09.29.510132