A conserved catalytic residue in the ubiquitin-conjugating enzyme family

Pei Ying Wu, Mary Hanlon, Michael Eddins, Colleen Tsui, Richard S. Rogers, Jane P. Jensen, Michael J. Matunis, Allan M. Weissman, Cynthia P. Wolberger, Cecile M. Pickart

Research output: Contribution to journalArticle

Abstract

Ubiquitin (Ub) regulates diverse functions in eukaryotes through its attachment to other proteins. The defining step in this protein modification pathway is the attack of a substrate lysine residue on Ub bound through its C-terminus to the active site cysteine residue of a Ub-conjugating enzyme (E2) or certain Ub ligases (E3s). So far, these E2 and E3 cysteine residues are the only enzyme groups known to participate in the catalysis of conjugation. Here we show that a strictly conserved E2 asparagine residue is critical for catalysis of E2- and E2/RING E3-dependent isopeptide bond formation, but dispensable for upstream and downstream reactions of Ub thiol ester formation. In constrast, the strictly conserved histidine and proline residues immediately upstream of the asparagine are dispensable for catalysis of isopeptide bond formation. We propose that the conserved asparagine side chain stabilizes the oxyanion intermediate formed during lysine attack. The E2 asparagine is the first non-covalent catalytic group to be proposed in any Ub conjugation factor.

Original languageEnglish (US)
Pages (from-to)5241-5250
Number of pages10
JournalEMBO Journal
Volume22
Issue number19
DOIs
StatePublished - Oct 1 2003

Keywords

  • Catalytic mechanism
  • E2
  • E3
  • Isopeptide
  • Ubiquitin

ASJC Scopus subject areas

  • Neuroscience(all)
  • Molecular Biology
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

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  • Cite this

    Wu, P. Y., Hanlon, M., Eddins, M., Tsui, C., Rogers, R. S., Jensen, J. P., Matunis, M. J., Weissman, A. M., Wolberger, C. P., & Pickart, C. M. (2003). A conserved catalytic residue in the ubiquitin-conjugating enzyme family. EMBO Journal, 22(19), 5241-5250. https://doi.org/10.1093/emboj/cdg501