title: Direct catalysis of lysine 48-linked polyubiquitin chains by the ubiquitin-activating enzyme.
abstract: Within the ubiquitin degradation pathway, the canonical signal is a lysine 48-linked polyubiquitin chain that is assembled upon an internal lysine residue of a substrate protein. Once constructed, this ubiquitin chain becomes the principle signal for recognition and target degradation by the 26S proteasome. The mechanism by which polyubiquitin chains are assembled on a substrate protein, however, has yet to be clearly defined. In an in vitro model system, purified E2-ubiquitin thiolester was unable to catalyze the formation of polyubiquitin chains in the absence of the ubiquitin-activating enzyme E1. Mutagenesis of key residues within the E1 active site revealed that its conserved catalytic cysteine residue is essential for the formation of these chains. Moreover, inactivation of the E2 active site had no effect on the ability of E1 to catalyze ubiquitin chain formation. These findings strongly suggest E1 is responsible for not only the activation of ubiquitin but also for the direct catalytic extension of a lysine 48-linked polyubiquitin chain.
authors.affiliation: Department of Biochemistry and Institute for Biomolecular Design, University of Alberta, Edmonton, Alberta, Canada.
authors.forename: J Torin
authors.initials: JT
authors.lastname: Huzil
authors.forename: Rajeet
authors.initials: R
authors.lastname: Pannu
authors.forename: Christopher
authors.initials: C
authors.lastname: Ptak
authors.forename: Grace
authors.initials: G
authors.lastname: Garen
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Ellison
mesh_terms: D001665:Binding Sites; D002384:Catalysis; D004926:Escherichia coli / Q000201:enzymology / Q000378:metabolism; D008025:Ligases / Q000737:chemistry; D008239:Lysine / Q000737:chemistry*; D016296:Mutagenesis; D025821:Polyubiquitin / Q000737:chemistry*; D011499:Protein Processing, Post-Translational; D025801:Ubiquitin / Q000737:chemistry; D044764:Ubiquitin-Activating Enzymes / Q000737:chemistry*; D044763:Ubiquitin-Conjugating Enzymes / Q000737:chemistry
chemical_list: D025801:Ubiquitin; D025821:Polyubiquitin; D044763:Ubiquitin-Conjugating Enzymes; D008025:Ligases; D044764:Ubiquitin-Activating Enzymes; D008239:Lysine