Affordable Access

deepdyve-link
Publisher Website

Detection and characterization of SUMO protease activity using a sensitive enzyme-based reporter assay.

Authors
  • Leach, Craig A
  • Tian, Xufan
  • Mattern, Michael R
  • Nicholson, Benjamin
Type
Published Article
Journal
Methods in Molecular Biology
Publication Date
Jan 01, 2009
Volume
497
Pages
269–281
Identifiers
DOI: 10.1007/978-1-59745-566-4_18
PMID: 19107424
Source
Medline
License
Unknown

Abstract

In this chapter we describe a novel, sensitive, homogenous high throughput reporter-based in vitro assay for SUMO protease activity developed by Progenra, Inc. A reporter construct was created by fusing His(6)-tagged small ubiquitin-like modifier (SUMO) to the amino terminus of the reporter enzyme phospholipase A(2) (PLA(2)). Following cleavage by a member of the sentrin specific proteases (SENPs), free PLA(2) is able to turn over its substrate, resulting in the release of a fluorescent product which is readily quantifiable using a fluorimeter or a fluorescence plate reader. The utility of this SUMO-CHOP-Reporter assay platform is demonstrated by its ability to determine K(m) values and to characterize inhibitors of SUMO proteases.

Report this publication

Statistics

Seen <100 times