The TRAF6 RING finger domain mediates physical interaction with Ubc13.
Tumor necrosis factor receptor associated factor 6 (TRAF6) is an important signaling molecule involved in a diverse array of physiological processes. It has been proposed that TRAF6, a RING finger-containing protein, acts as a ubiquitin ligase (E3) and a target for Lys-63 linked polyubiquitination mediated by Ubc13-Uev, a ubiquitin conjugating (E2) complex. However, the physical interaction between TRAF6 and this E2 complex has not been reported. We used the yeast two-hybrid assay to demonstrate that TRAF6 indeed interacts with the E2 complex through its direct binding to Ubc13. Either a single Cys-to-Ser substitution within the TRAF6 RING finger domain or an amino acid substitution on the Ubc13 surface, that is predicted to interact with RING finger proteins, is able to abolish the interaction. In addition, we found that TRAF6 can interact with itself and this self-interaction domain is mapped to the N-terminus containing the RING finger motif. Based on this study and our previous Ubc13-Uev structural analysis, the interface of Ubc13-TRAF6 RING finger can be predicted.
Wooff
Jill
J
Department of Microbiology and Immunology, University of Saskatchewan, Saskatoon, SK, Canada S7N 5E5.
Pastushok
Landon
L
Hanna
Michelle
M
Fu
Yu
Y
Xiao
Wei
W
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D000818:Animals; D017353:Gene Deletion; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011506:Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000235:genetics; D048029:TNF Receptor-Associated Factor 6; D014162:Transfection; D014170:Transformation, Genetic; D020798:Two-Hybrid System Techniques; D044763:Ubiquitin-Conjugating Enzymes / Q000235:genetics / Q000378:metabolism*
D011506:Proteins; D011993:Recombinant Fusion Proteins; D048029:TNF Receptor-Associated Factor 6; C471607:UBE2N protein, human; D044763:Ubiquitin-Conjugating Enzymes