title: A parallel affinity purification method for selective isolation of polyubiquitinated proteins.
abstract: We developed a parallel affinity purification (PAP) procedure, in which ubiquitinated proteins are purified from the cells that coexpress two affinity-tagged ubiquitins by sequential use of affinity chromatography specific to each tag. In contrast with previous procedures using a single affinity-tagged ubiquitin, the PAP eliminates highly abundant ubiquitin monomers and monoubiquitinated proteins to selectively enrich proteins bearing both affinity-tags, or poly- and multiubiquitinated proteins. Accordingly, it would serve as a powerful method to facilitate mass-spectrometric identification of ubiquitinated proteins.
authors.affiliation: Department of Computational Biology, Graduate School of Frontier Sciences, University of Tokyo, Kashiwa, Japan.
authors.forename: Kazuhisa
authors.initials: K
authors.lastname: Ota
authors.forename: Keiji
authors.initials: K
authors.lastname: Kito
authors.forename: Shun-ichiro
authors.initials: S
authors.lastname: Iemura
authors.forename: Tohru
authors.initials: T
authors.lastname: Natsume
authors.forename: Takashi
authors.initials: T
authors.lastname: Ito
mesh_terms: D002846:Chromatography, Affinity / Q000379:methods*; D025821:Polyubiquitin / Q000378:metabolism*; D011506:Proteins / Q000302:isolation & purification* / Q000378:metabolism; D040901:Proteomics / Q000379:methods*; D015203:Reproducibility of Results; D054875:Ubiquitination
chemical_list: D011506:Proteins; D025821:Polyubiquitin