Monoubiquitylation of GGA3 by hVPS18 regulates its ubiquitin-binding ability.
GGAs (Golgi-localizing, gamma-adaptin ear domain homology, ADP-ribosylation factor (ARF)-binding proteins), constitute a family of monomeric adaptor proteins and are associated with protein trafficking from the trans-Golgi network to endosomes. Here, we show that GGA3 is monoubiquitylated by a RING-H2 type-ubiquitin ligase hVPS18 (human homologue of vacuolar protein sorting 18). By ... in vitro ubiquitylation assays, we have identified lysine 258 in the GAT domain as a major ubiquitylation site that resides adjacent to the ubiquitin-binding site. The ubiquitylation is abolished by a mutation in either the GAT domain or ubiquitin that disrupts the GAT-ubiquitin interaction, indicating that the ubiquitin binding is a prerequisite for the ubiquitylation. Furthermore, the GAT domain ubiquitylated by hVPS18 no longer binds to ubiquitin, indicating that ubiquitylation negatively regulates the ubiquitin-binding ability of the GAT domain. These results suggest that the ubiquitin binding and ubiquitylation of GGA3-GAT domain are mutually inseparable through a ubiquitin ligase activity of hVPS18.
Mesh Terms:
ADP-Ribosylation Factors, Adaptor Proteins, Vesicular Transport, Binding Sites, Crystallography, X-Ray, Hela Cells, Humans, Lysine, Models, Molecular, Protein Binding, Protein Structure, Quaternary, Ubiquitin, Ubiquitin-Protein Ligases, Vesicular Transport Proteins
ADP-Ribosylation Factors, Adaptor Proteins, Vesicular Transport, Binding Sites, Crystallography, X-Ray, Hela Cells, Humans, Lysine, Models, Molecular, Protein Binding, Protein Structure, Quaternary, Ubiquitin, Ubiquitin-Protein Ligases, Vesicular Transport Proteins
Biochem. Biophys. Res. Commun.
Date: Nov. 10, 2006
PubMed ID: 16996030
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