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In vitro analysis of proteasome-associated USP14 activity for substrate degradation and deubiquitylation

In vitro analysis of proteasome-associated USP14 activity for substrate degradation and deubiquitylation
Muniyappan, SrinivasanLee, Byung-Hoon
DGIST Authors
Muniyappan, SrinivasanLee, Byung-Hoon
Issued Date
Article Type
Review; Book Chapter
Author Keywords
Deubiquitinating enzymeIn vitro deubiquitination assayProteasomeUbiquitin adductsUbiquitin conjugateUSP14
The ubiquitin-proteasome pathway plays an essential role in maintaining protein homeostasis and regulates almost every aspect of cellular processes in eukaryotes. Emerging evidence indicates that the proteasome does not work as a simple unidirectional molecular machinery for substrate proteolysis. In fact, proteasome activity should be tightly regulated, and the proteasome itself can be dynamically engaged in the degradation cycle. Proteasome-mediated degradation can occur through multistep mechanisms such as ubiquitin-dependent substrate recognition, deubiquitination, and ATP-driven unfolding and translocation of the substrate into 20S chamber for proteolytic cleavage. Deubiquitination is particularly interesting because this reaction may impose a critical checkpoint for substrate turnover on the proteasome. Notably, there are three major deubiquitinating enzymes (DUBs) on human proteasomes: USP14, UCH37, and RPN11. USP14 can spare the substrate from degradation prior to the proteasome's commitment step, suggesting that USP14 inhibition may stimulate proteasomal degradation of undesirable proteins under certain proteotoxic conditions. Furthermore, USP14 deubiquitinates multichain conjugates, the first among ~ 100 DUBs found to have this striking specificity. In this chapter, we describe in vitro methods to test proteasome-associated USP14 activity for substrate degradation and deubiquitylation. © 2019 Elsevier Inc.
Academic Press
Related Researcher
  • 이병훈 Lee, Byung-Hoon 뉴바이올로지학과
  • Research Interests Ubiquitin-proteasome system; Protein homeostasis; Small-molecule chemical screening and drug discovery in human disease
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Department of New Biology Lab of Protein Homeostasis and Drug Discovery 1. Journal Articles


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