A time-resolved fluorescence resonance energy transfer-based assay for DEN1 peptidase activity

被引:13
|
作者
Engels, Ingo H. [1 ]
Daguia, Catherine [1 ]
Huynh, Truc [1 ]
Urbina, Hugo [1 ]
Buddenkotte, Joerg [1 ]
Schumacher, Andrew [1 ]
Caldwell, Jeremy S. [1 ]
Brinker, Achim [1 ]
机构
[1] Genom Inst Novartis Res Fdn, San Diego, CA 92121 USA
关键词
UBIQUITIN-LIKE PROTEIN; NEDD8; ENZYMES; FAMILY; NEDP1; CUL1;
D O I
10.1016/j.ab.2009.03.035
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
Neural precursor cell expressed, developmentally clown-regulated gene 8 (NEDD8) is a recently discovered ubiquitin-like posttranslational modifier. NEDD8 acts predominantly as a regulator of ubiquitin-protein ligases and as a decoy for proteins targeted for proteasomal degradation. It thereby controls key events in cell cycle Progression and embryogenesis. Deneddylase-1 (DEN1/NEDP1/SENP8) features a selective peptidase activity converting the proNEDD8 precursor to its mature form and an isopeptidase activity deconjugating NEDD8 from substrates Such as cullins and p53. In this study, we describe a high-throughput screening (HTS)-compatible time-resolved fluorescent resonance energy transfer (TR-FRET) assay measuring the peptidase activity of DEN1. (C) 2009 Elsevier Inc. All rights reserved.
引用
收藏
页码:85 / 87
页数:3
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