title: S5a/Rpn10, a UIM-protein, as a universal substrate for ubiquitination.
abstract: The assay of the activity of ubiquitin (Ub) ligases (E3s) and screens for pharmacological agents that alter their function are a continual challenge for basic investigators as well as in drug development. The assay of different E3s requires distinct detection methods and reagents (e.g., specific antibodies against each E3 or substrate). So, a single assay applicable to many E3s could be very useful. Here, we demonstrate that S5a/Rpn10 binds to the growing polyUb chain formed on a substrate (or on the Ub ligase during autoubiquitination) and then itself becomes extensively ubiquitinated. S5a thus can serve as a universal substrate for ubiquitination. This biochemical property of S5a provides a method for measuring the enzymatic activity of any E3. This approach is valuable when substrates are not known or not available and when multiple ubiquitination reactions are being studied (e.g., in high-throughput screens).
authors.affiliation: Department of Cell Biology, Harvard Medical School, Boston, MA, USA.
authors.forename: Hyoung Tae
authors.initials: HT
authors.lastname: Kim
authors.forename: Alfred L
authors.initials: AL
authors.lastname: Goldberg
mesh_terms: D006801:Humans; D025821:Polyubiquitin / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism*; D016601:RNA-Binding Proteins; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry / Q000378:metabolism; D054875:Ubiquitination
chemical_list: C498506:PSMD4 protein, human; D016601:RNA-Binding Proteins; D025821:Polyubiquitin; D044767:Ubiquitin-Protein Ligases; D046988:Proteasome Endopeptidase Complex; C061553:ATP dependent 26S protease