title: In vitro cleavage of Nedd8 from cullin 1 by COP9 signalosome and deneddylase 1.
abstract: Enzymatic cleavage of Nedd8 from its cullin conjugates plays a critical role in ubiquitin-dependent proteolysis by regulating the activity of the cullin-based RING-H2 E3 ubiquitin ligases. This chapter provides methods for the preparation of two Nedd8 isopeptidases: the COP9 signalosome and human deneddylase 1. It also describes the development of cell-free systems for cleavage of the Nedd8-cullin 1 isopeptide bond formed in vitro or in vivo.
authors.affiliation: Department of Oncological Sciences, Mount Sinai School of Medicine, New York, NY 10029-6574, USA.
authors.forename: Kosj
authors.initials: K
authors.lastname: Yamoah
authors.forename: Kenneth
authors.initials: K
authors.lastname: Wu
authors.forename: Zhen-Qiang
authors.initials: ZQ
authors.lastname: Pan
mesh_terms: D000075686:COP9 Signalosome Complex; D002352:Carrier Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D018797:Cell Cycle Proteins / Q000378:metabolism*; D002460:Cell Line; D002850:Chromatography, Gel / Q000379:methods; D044842:Cullin Proteins / Q000378:metabolism*; D010450:Endopeptidases / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D046912:Multiprotein Complexes / Q000302:isolation & purification / Q000378:metabolism*; D000075687:NEDD8 Protein; D009842:Oligopeptides; D010447:Peptide Hydrolases / Q000302:isolation & purification / Q000378:metabolism*; D010455:Peptides / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D010761:Phosphorus Radioisotopes; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D014452:Ubiquitins / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D018797:Cell Cycle Proteins; C100318:Cullin 1; D044842:Cullin Proteins; D046912:Multiprotein Complexes; D000075687:NEDD8 Protein; C082860:NEDD8 protein, human; D009842:Oligopeptides; D010455:Peptides; D010761:Phosphorus Radioisotopes; C479529:RBX1 protein, human; D011993:Recombinant Fusion Proteins; D014452:Ubiquitins; C085706:FLAG peptide; D010450:Endopeptidases; D010447:Peptide Hydrolases; D000075686:COP9 Signalosome Complex; C477008:SENP8 protein, human