title: E2-c-Cbl recognition is necessary but not sufficient for ubiquitination activity.
abstract: The E2 ubiquitin-conjugating enzymes UbcH7 and UbcH5B both show specific binding to the RING (really interesting new gene) domain of the E3 ubiquitin-protein ligase c-Cbl, but UbcH7 hardly supports ubiquitination of c-Cbl and substrate in a reconstituted system. Here, we found that neither structural changes nor subtle differences in the E2-E3 interaction surface are possible explanations for the functional specificity of UbcH5B and UbcH7 in their interaction with c-Cbl. The quick transfer of ubiquitin from the UbcH5B-Ub thioester to c-Cbl or other ubiquitin acceptors suggests that UbcH5B might functionally be a relatively pliable E2 enzyme. In contrast, the UbcH7-Ub thioester is too stable to transfer ubiquitin under our assay conditions, indicating that UbcH7 might be a more specific E2 enzyme. Our results imply that the interaction specificity between c-Cbl and E2 is required but not sufficient for transfer of ubiquitin to potential targets.
authors.affiliation: Department of NMR Spectroscopy, Bijvoet Center for Biomolecular Research, Utrecht University, Padualaan 8, 3584 CH, Utrecht, The Netherlands.
authors.forename: Anding
authors.initials: A
authors.lastname: Huang
authors.forename: Rob N
authors.initials: RN
authors.lastname: de Jong
authors.forename: Hans
authors.initials: H
authors.lastname: Wienk
authors.forename: G Sebastiaan
authors.initials: GS
authors.lastname: Winkler
authors.forename: H Th Marc
authors.initials: HT
authors.lastname: Timmers
authors.forename: Rolf
authors.initials: R
authors.lastname: Boelens
mesh_terms: D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D020836:Protein Structure, Quaternary; D017434:Protein Structure, Tertiary; D050721:Proto-Oncogene Proteins c-cbl / Q000235:genetics / Q000378:metabolism*; D044763:Ubiquitin-Conjugating Enzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D054875:Ubiquitination
chemical_list: C097309:UBE2D2 protein, human; C079437:UBE2L3 protein, human; D044763:Ubiquitin-Conjugating Enzymes; D050721:Proto-Oncogene Proteins c-cbl