Multifaceted N-Degron Recognition and Ubiquitylation by GID/CTLH E3 Ligases

N-degron E3 ubiquitin ligases recognize specific residues at the N-termini of substrates. Although molecular details of N-degron recognition are known for several E3 ligases, the range of N-terminal motifs that can bind a given E3 substrate binding domain remains unclear. Here, we discovered capacity of Gid4 and Gid10 substrate receptor subunits of yeast GID/human CTLH multiprotein E3 ligases to tightly bind a wide range of N-terminal residues whose recognition is determined in part by the downstream sequence context. Screening of phage displaying peptide libraries with exposed N-termini identified novel consensus motifs with non-Pro N-terminal residues binding Gid4 or Gid10 with high affinity. Structural data reveal that conformations of flexible loops in Gid4 and Gid10 complement sequences and folds of interacting peptides. Together with analysis of endogenous substrate degrons, the data show that degron identity, substrate domains harboring targeted lysines, and varying E3 ligase higher-order assemblies combinatorially determine efficiency of ubiquitylation and degradation. (c) 2021 The Author(s). Published by Elsevier Ltd. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/).

Automated summary

The study identified the binding capacity of Gid4 and Gid10 substrate receptor subunits to a wide range of N-terminal residues, influenced by the downstream sequence context. Screening of phage displaying peptide libraries revealed novel consensus motifs with high affinity binding to Gid4 and Gid10, with structural data showing the complementary conformations with interacting peptides. The research demonstrates that degron identity, substrate domains containing targeted lysines, and the composition of E3 ligase higher-order assemblies collectively determine the efficiency of ubiquitylation and degradation.