Structure of the ubiquitin hydrolase UCH-L3 complexed with a suicide substrate

Shahram Misaghi, Paul J. Galardy, Wim J.N. Meester, Huib Ovaa, Hidde L. Ploegh, Rachelle Gaudet

Research output: Contribution to journalArticlepeer-review

149 Scopus citations


Ubiquitin C-terminal hydrolases (UCHs) comprise a family of small ubiquitin-specific proteases of uncertain function. Although no cellular substrates have been identified for UCHs, their highly tissue-specific expression patterns and the association of UCH-L1 mutations with human disease strongly suggest a critical role. The structure of the yeast UCH Yuh1-ubiquitin aldehyde complex identified an active site crossover loop predicted to limit the size of suitable substrates. We report the 1.45 Å resolution crystal structure of human UCH-L3 in complex with the inhibitor ubiquitin vinylmethylester, an inhibitor that forms a covalent adduct with the active site cysteine of ubiquitin-specific proteases. This structure confirms the predicted mechanism of the inhibitor and allows the direct comparison of a UCH family enzyme in the free and ligand-bound state. We also show the efficient hydrolysis by human UCH-L3 of a 13-residue peptide in isopeptide linkage with ubiquitin, consistent with considerable flexibility in UCH substrate size. We propose a model for the catalytic cycle of UCH family members which accounts for the hydrolysis of larger ubiquitin conjugates.

Original languageEnglish (US)
Pages (from-to)1512-1520
Number of pages9
JournalJournal of Biological Chemistry
Issue number2
StatePublished - Jan 14 2005

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology


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