E3 ligase activity of RING finger proteins that interact with Hip-2, a human ubiquitin-conjugating enzyme

To identify proteins that interact with Huntingtin-interacting protein-2 (H ip-2), a ubiquitin-conjugating enzyme, a yeast two-hybrid screen system was used to isolate five positive clones. Sequence analyses showed that, with one exception, all Hip-2-interacting proteins Contained the RING finger mot ifs. The interaction of Hip-2 with RNF2, one of the clones, was further con firmed through in vitro and in vivo experiments. Mutations in the RING doma in of RNF2 prevented the clone from binding to Hip-2, an indication that th e RING domain is the binding determinant. RNF2 showed a ubiquitin ligase (E 3) activity in the presence of Hip-2, suggesting that a subset of RING fing er proteins may have roles as Us. (C) 2001 Published by Elsevier Science B. V. on behalf of the Federation of European Biochemical Societies.