Poster Presentation Inaugural Australian Ubiquitin Summit 2025

TRIP12 structures reveal mechanisms underlying HECT E3-mediated formation of K29 linkages and branched ubiquitin chains (#116)

Samuel A. Maiwald 1 , Laura A. Schneider 1 , Ronnald Vollrath 1 , Joanna Liwocha 1 , Matthew D. Maletic 2 , Kirby N. Swatek 1 , Monique P. C. Mulder 2 , Brenda A. Schulman 1
  1. Department of Molecular Machines and Signaling, Max Planck Institute of Biochemistry, Martinsried, Bavaria, Germany
  2. Department of Cell and Chemical Biology, Leiden University Medical Center, Leiden, Zuid-Holland, Netherlands

Precise regulation by the ubiquitin system hinges on E3 ligases assembling ubiquitin chains with specific topologies1, but the mechanisms underlying the formation of atypical linkages remain poorly understood. We employed biochemistry, chemical biology, and cryo-EM to elucidate how the human HECT E3 ligase TRIP12 catalyzes the formation of K29-linked and K29/K48-branched ubiquitin chains. TRIP12 adopts a pincer-like architecture: one side consists of tandem ubiquitin-binding domains that orient the proximal ubiquitin to position K29 in the active site, and selectively engage a distal ubiquitin from a K48-linked chain. The opposite side, comprising the HECT domain, precisely juxtaposes the donor and acceptor ubiquitins to enforce K29 linkage specificity. Structural comparison with the K48-specific HECT E3 ligase UBR5 uncovers a shared mechanism: conserved features of the E3 ligases, donor, and acceptor ubiquitins configure the active site, while E3-specific domains support acceptor positioning to enable linkage-specific chain formation.

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