Premium
A parallel affinity purification method for selective isolation of polyubiquitinated proteins
Author(s) -
Ota Kazuhisa,
Kito Keiji,
Iemura Shunichiro,
Natsume Tohru,
Ito Takashi
Publication year - 2008
Publication title -
proteomics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.26
H-Index - 167
eISSN - 1615-9861
pISSN - 1615-9853
DOI - 10.1002/pmic.200800271
Subject(s) - ubiquitins , affinity chromatography , ubiquitin , chemistry , biochemistry , tandem affinity purification , affinity electrophoresis , chromatography , target protein , ubiquitin ligase , enzyme , gene
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.