Specific and covalent targeting of conjugating and deconjugating enzymes of ubiquitin-like proteins.

ABSTRACT: Modification of proteins by ubiquitin (Ub)-like proteins (UBLs) plays an important role in many cellular processes, including cell cycle progression, nuclear transport, and autophagy. Protein modification occurs via UBL-conjugating and -deconjugating enzymes, which presumably exert a regulatory function by determining the conjugation status of the substrate proteins. To target and identify UBL-modifying enzymes, we produced Nedd8, ISG15, and SUMO-1 in Escherichia coli and equipped them with a C-terminal electrophilic trap (vinyl sulfone [VS]) via an intein-based method. These C-terminally modified UBL probes reacted with purified UBL-activating (E1), -conjugating (E2), and -deconjugating enzymes in a covalent fashion. Modified UBLs were radioiodinated and incubated with cell lysates prepar

SUBMITTER: Hemelaar J 

PROVIDER: S-EPMC303361 | biostudies-literature | 2004 Jan

REPOSITORIES: biostudies-literature