title: Catalytically Important Residues of E6AP Ubiquitin Ligase Identified Using Acid-Cleavable Photo-Cross-Linkers.
abstract: Inactivation of the E6AP E3 ubiquitin ligase (UBE3A gene) causes Angelman syndrome, while aberrant degradation of p53 by E6AP is implicated in cervical cancers. Herein, we describe the development of photo-cross-linkers to discover catalytic residues of E6AP. Using these cross-linkers, we identified covalent modifications of the E6AP catalytic cysteine and two lysines: Lys(847) and Lys(799). Lys(847) is required for the formation of Lys(48)-linked polyubiquitin chains, while the K799A E6AP mutant was more active at producing Lys(48)-linked polyubiquitin chains. Thus, opposing roles of Lys(799) and Lys(847) pave the path forward to pharmacological inhibitors or activators of E6AP for therapeutic purposes.
authors.affiliation: Chemistry of Life Processes Institute, Department of Chemistry, Northwestern University, Silverman Hall, 2145 Sheridan Road, Evanston, Illinois 60208, United States.
authors.forename: David T
authors.initials: DT
authors.lastname: Krist
authors.affiliation: Chemistry of Life Processes Institute, Department of Chemistry, Northwestern University, Silverman Hall, 2145 Sheridan Road, Evanston, Illinois 60208, United States.
authors.forename: Alexander V
authors.initials: AV
authors.lastname: Statsyuk
mesh_terms: D019943:Amino Acid Substitution; D020134:Catalytic Domain; D003432:Cross-Linking Reagents / Q000737:chemistry*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D055668:Photochemical Processes; D025821:Polyubiquitin / Q000502:physiology; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry* / Q000235:genetics; D054875:Ubiquitination
chemical_list: D003432:Cross-Linking Reagents; D025821:Polyubiquitin; C471639:UBE3A protein, human; D044767:Ubiquitin-Protein Ligases