title: Large-scale analysis of the human ubiquitin-related proteome.
abstract: Protein ubiquitylation contributes to the regulation of many cellular processes including protein degradation, receptor internalization, and repair of DNA damage. We now present a comprehensive characterization of ubiquitin-conjugated and ubiquitin-associated proteins in human cells. The proteins were purified by immunoaffinity chromatography under denaturing or native conditions. They were then digested with trypsin, and the resulting peptides were analyzed by 2-D LC and MS/MS. A total of 670 distinct proteins were identified; 345 proteins (51%) were classified as Urp-D (ubiquitin-related proteome under the denaturing condition) and comprised ubiquitin-conjugated molecules, whereas 325 proteins (49%) were classified as Urp-N (ubiquitin-related proteome only under the native condition) and included molecules that associated with ubiquitylated proteins. The proportions of proteins in various functional categories differed substantially between Urp-D and Urp-N. Many ribosomal subunits were detected in the Urp-D group of proteins and several of these subunits were directly shown to be ubiquitylated by mass spectrometric analysis, suggesting that ubiquitylation might play an important role in the regulation and/or quality control of ribosomal proteins. Our results demonstrate the potential of proteomics analysis of protein ubiquitylation to provide important insight into the regulation of protein stability and other ubiquitin-related cellular functions.
authors.affiliation: Department of Molecular and Cellular Biology, Medical Institute of Bioregulation, Kyushu University, Fukuoka, Fukuoka, Japan.
authors.forename: Masaki
authors.initials: M
authors.lastname: Matsumoto
authors.forename: Shigetsugu
authors.initials: S
authors.lastname: Hatakeyama
authors.forename: Koji
authors.initials: K
authors.lastname: Oyamada
authors.forename: Yoshiya
authors.initials: Y
authors.lastname: Oda
authors.forename: Toshihide
authors.initials: T
authors.lastname: Nishimura
authors.forename: Keiichi I
authors.initials: KI
authors.lastname: Nakayama
mesh_terms: D001665:Binding Sites; D002460:Cell Line; D002846:Chromatography, Affinity; D002853:Chromatography, Liquid; D006801:Humans; D013058:Mass Spectrometry; D011485:Protein Binding; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D020543:Proteome / Q000032:analysis* / Q000378:metabolism; D012270:Ribosomes / Q000737:chemistry / Q000378:metabolism; D025801:Ubiquitin / Q000032:analysis* / Q000378:metabolism
chemical_list: D021122:Protein Subunits; D020543:Proteome; D025801:Ubiquitin