E1-catalyzed ubiquitin C-terminal amidation for the facile synthesis of deubiquitinase substrates

Chembiochem. 2014 Jan 3;15(1):37-41. doi: 10.1002/cbic.201300608. Epub 2013 Dec 16.

Abstract

Will Ub my partner? The ubiquitin (Ub)-activating enzyme (E1) was used to catalyze an amidation reaction to functionalize the C terminus of Ub with unique functional groups, such as thiol, azide, alkyne, and alkene groups, with high efficiency and yield (>90 %). These groups were then applied for the facile synthesis of fluorophore-conjugated ubiquitin and specifically conjugated diubiquitin substrates for deubiquitinases.

Keywords: deubiquitinases; diubiquitin; protein engineering; synthetic biology; ubiquitin functionalization.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alkenes / chemistry
  • Alkenes / metabolism
  • Alkynes / chemistry
  • Alkynes / metabolism
  • Animals
  • Azides / chemistry
  • Azides / metabolism
  • Biocatalysis
  • Kinetics
  • Protein Structure, Tertiary
  • Substrate Specificity
  • Sulfhydryl Compounds / chemistry
  • Sulfhydryl Compounds / metabolism
  • Ubiquitin / chemistry
  • Ubiquitin / metabolism*
  • Ubiquitin-Activating Enzymes / chemistry
  • Ubiquitin-Activating Enzymes / metabolism*
  • Ubiquitin-Specific Proteases / chemistry
  • Ubiquitin-Specific Proteases / metabolism*
  • Ubiquitination

Substances

  • Alkenes
  • Alkynes
  • Azides
  • Sulfhydryl Compounds
  • Ubiquitin
  • Ubiquitin-Specific Proteases
  • Ubiquitin-Activating Enzymes