The differential modulation of USP activity by internal regulatory domains, interactors and eight ubiquitin chain types

Chem Biol. 2011 Dec 23;18(12):1550-61. doi: 10.1016/j.chembiol.2011.10.017.


Ubiquitin-specific proteases (USPs) are papain-like isopeptidases with variable inter- and intramolecular regulatory domains. To understand the effect of these domains on USP activity, we have analyzed the enzyme kinetics of 12 USPs in the presence and absence of modulators using synthetic reagents. This revealed variations of several orders of magnitude in both the catalytic turnover (k(cat)) and ubiquitin (Ub) binding (K(M)) between USPs. Further activity modulation by intramolecular domains affects both the k(cat) and K(M), whereas the intermolecular activators UAF1 and GMPS mainly increase the k(cat). Also, we provide the first comprehensive analysis comparing Ub chain preference. USPs can hydrolyze all linkages and show modest Ub-chain preferences, although some show a lack of activity toward linear di-Ub. This comprehensive kinetic analysis highlights the variability within the USP family.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Catalytic Domain
  • Endopeptidases / chemistry
  • Endopeptidases / genetics
  • Endopeptidases / metabolism*
  • Guanosine Monophosphate / metabolism
  • Humans
  • Kinetics
  • Nuclear Proteins / metabolism
  • Protein Binding
  • Protein Structure, Tertiary
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Thionucleotides / metabolism
  • Ubiquitin / chemistry
  • Ubiquitin / metabolism*
  • Ubiquitin-Specific Proteases


  • Nuclear Proteins
  • Recombinant Proteins
  • Thionucleotides
  • USP1 associated factor 1, human
  • Ubiquitin
  • guanosine 5'-monophosphorothioate
  • Guanosine Monophosphate
  • Endopeptidases
  • Ubiquitin-Specific Proteases