title: Identification of phosphorylation sites on the E3 ubiquitin ligase UBR5/EDD.
abstract: UBR5 (ubiquitin protein ligase E3 component n-recognin 5)/EDD (E3 ligase identified by differential display) is an E3 ubiquitin ligase that is a potential biomarker for poor prognosis for recurrent, platinum-resistant ovarian cancer. UBR5 has a role in the DNA damage response and many such proteins are regulated by phosphorylation. UBR5 is a 309 kDa nuclear phosphoprotein that we previously identified as a substrate of the MAP kinase ERK2. With its 477 potential phosphorylation sites, little is known about UBR5 phosphorylation and how it may regulate protein function. Currently, thirty-four sites of phosphorylation on UBR5 have been reported in the literature, mostly identified by large scale proteomics studies of tissues or of cells after various treatments; however, no studies have specifically targeted the identification of UBR5 phosphorylation sites. In this study, we used Liquid Chromatography-Mass Spectrometry (LC-MS/MS) to obtain a total sequence coverage of 64.3% from combining tryptic and GluC digests on UBR5 isolated from transfected COS-1 cells. We identified 24 sites of phosphorylation, 18 of which are novel sites. This data enhances our knowledge of UBR5 phosphorylation and provides a framework for the study of how phosphorylation affects UBR5 function.
authors.affiliation: Department of Cell and Molecular Pharmacology and Experimental Therapeutics, Medical University of South Carolina, Charleston, SC 29425, USA.
authors.forename: Jennifer Rutherford
authors.initials: JR
authors.lastname: Bethard
authors.forename: Hui
authors.initials: H
authors.lastname: Zheng
authors.forename: Lawton
authors.initials: L
authors.lastname: Roberts
authors.forename: Scott T
authors.initials: ST
authors.lastname: Eblen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D002853:Chromatography, Liquid; D010766:Phosphorylation; D053719:Tandem Mass Spectrometry; D044767:Ubiquitin-Protein Ligases / Q000737:chemistry / Q000378:metabolism*
chemical_list: C117617:UBR5 protein, human; D044767:Ubiquitin-Protein Ligases